2^nd International Conference on Systems and Synthetic Biology August 18-20, 2016 London, UK

Biography:

Dr. Jean A. Boutin graduated (Thèse d’Etat en Sciences Biologiques) from Nancy University (France) on drug metabolism. He did his postdoctoral training at Johns Hopkins (Baltimore) and at the Karolinska Institutet (Stockholm, Sweden). He was hired as protein chemist in Les Laboratoires SERVIER (LLS) in 1986. During the 30ish last years, Dr. Boutin moved from oncology to peptide research and then molecular & cellular pharmacology. Recently, LLS created a drug discovery platform that Dr. Boutin leads. This structure involves all the aspects of drug discovery, from molecular modeling to ligand/protein biophysical interaction measurements, including protein chemistry, stem cells, structural biology, chemogenetics, HTS, biophysics, Biologics…The main interests of Dr. Boutin are N-myristoyltransferase, melatonin, quinone reductase 2, MCH and autotaxin. In relation with Biologics, our group more particularly explores all the areas related to the possibilities to incorporate exotic amino acids into proteins, especially enzymes.

Abstract:

Synthetic biology is a growing field in which the contribution brought by chemical synthesis of proteins and particularly enzymes is fundamental. Despite this fact, the chemical synthesis of catalytic active proteins remains poorly documented, essentially because it is hard to obtain enough material to use it in biochemical experiments. Chemical synthesis of proteins could permit to have access to the incorporation of unnatural (exotic) amino acids into catalytic active proteins, a feature amenable by recombinant technologies, but that requires delicate manipulation of the bacterial machinery. The developments brought by this approach, include but are not limited to the measure of the influence of unnatural (exotic) amino acids on the 3D structure of enzyme, its activity as well as their recognition of substrate, co-substrate or regulator. The main limitation remains a quantitative problem: How to progress from microgram of proteins produced nowadays with essentially recombinant techniques to tens of milligrams? We chose to present as a model the synthesis and thorough characterization of calstabin, a short protein proline isomerase of 107 amino acids. The protein was synthesized using the native chemical ligation approach. Several tens of milligrams were obtained. Therefore, we were able to refold the polypeptide properly, to characterize its biophysical properties, to measure its catalytic activity and finally to crystallize it in order to obtain its tridimensional structure after X-ray diffraction. Further to it and as a first step of validation of the whole process, we incorporated exotic amino acids in the easiest reachable part of the protein N-terminus. Avenues are now open to obtain further proteins modified with exotic amino acids in a way that is only barely accessible by molecular biology. We hope that these approaches will permit to gain detailed information on the structure-function relationship of proteins as long as they are reachable by complete chemical synthesis (below 300 amino acids).

Biography:

Roberto Mazzoli has completed his PhD in 2003 and he has been working as an Assistant Professor in Biochemistry at the University of Turin, Italy since 2011. He has been studying the metabolism of microorganisms aimed at industrial processes. His main research activity has concerned metabolic engineering of microorganisms for cellulosic consolidated biorefinery. He is the PI of the group of Proteomics and Metabolic Engineering of Prokaryotes at DBIOS, University of Turin. He has published 21 papers in peer-reviewed international journals, 1 book chapter and 2 international patents and he is serving as Editorial Board Member or Reviewer for several reputed journals.

Abstract:

Cellulosic biomass is the most interesting substrate for biorefinery, owing to its abundance and low cost. However, cellulose is highly recalcitrant to biodegradation. Currently available technologies for fermenting cellulose to industrially relevant compounds (e.g., fuels, plastics) consist of multiple-step processes which are not cost-competitive with traditional petrochemical refinery. Engineering recombinant microbial strains able to catalyze single-step fermentation (i.e., consolidated bioprocessing, CBP) of biomass into high-value products is a key goal so as to develop cost-competitive cellulosic biomass biorefinery. For at least 20 years, intensive research efforts have been dedicated to develop recombinant strains suitable for cellulose-CBP following either the “native cellulolytic strategy”, aimed at engineering natural cellulolytic microorganisms so as to improve their product formation features or the “recombinant cellulolytic strategy” aimed at conferring cellulolytic ability to strains which naturally produce industrially-relevant products with high efficiency. Although these researches have attained impressive results, current forefront achievements yet resemble more to proofs-of-concept than to new biotechnological deliverables ready for application in industrial fermentations. The present contribution aims at providing an overview on most significant successful examples of these strategies together with major issues which still need to be addressed and possible solutions.

Biography:

Taizo Uda has obtained his PhD from Kyushu University and Postdoctoral studies from Wisconsin University-Milwaukee. He has studied on heterogeneous catalysis and antibody engineering. He was the Director of Diagnostic Group of a Research Institute in a Chemical Company. He is a Visiting Professor at Oita University and Kyushu University. He has published more than 50 papers in reputed journals.

Abstract:

Issue on the structural diversity (heterogeneity) of the molecule has been focused along with the development of recombinant antibody drugs. The structural diversity provides some (or many) isoforms of an antibody caused by different charges, different molecular sizes and/or modifications of amino acid residues. For practical use, the antibody and/or the subunits must have a defined structure. A whole antibody is consisted of the light and heavy chain. Once they are separated, the structure of the light or heavy chain becomes very flexible, which also causes the structural diversity and then gives some kinds of isoforms of different pI. We prepared several human antibody light chains possessing a C-terminal histidine-tag, which was expressed in E. coli. After Ni-NTA chromatography, the purified light chain was subjected to the cation-exchange chromatography, where several peaks consisted of the monomers and/or dimers were observed at the different retention time. This suggests that the different forms in both molecular sizes and the electrical charges co-exist in the solution, while only a light chain is present. This was the similar results regarding molecular heterogeneity as those observed in recombinant antibody drugs as reported. Several metal ions were examined to investigate the effect on the structural heterogeneity (diversity). Note that copper ion exhibited huge effect for solving the heterogeneity (diversity) issue. In the presentation, the role of the constant domain will be also introduced in detail.

Biography:

Paul Lomax is Product Manager for TTP Labtech, sample management products. With over 15 years of experience in the automation of life science applications, he is responsible for TTP Labtech’s range of novel automated -20°C and -80°C storage systems.

Abstract:

With momentum building in the establishment of Synthetic biology foundries, it is evident that automation plays a key role throughout the Synthetic Biology workflow. Safe, secure and robust inventory storage and management is critical. To maximize efficient operation, rapid access to stock constructs or “biobricks” is key and TTP Labtech offer solutions for automated storage at -20°C and -80°C with true cherry picking ensuring that only the required samples are retrieved. 2D barcoded labware allows samples to be tracked and stored and retrieved with the simplicity and speed of a vending machine. Novel pneumatic technology to move tubes minimizes the need for robotics in the cold zone, providing long term reliability of operation. Learn more about these how TTP Labtech’s solutions could automate the heart of your Synthetic Biology facility. Paul Lomax is a global product manager for TTP Labtech, who are a UK based developer and supplier of automation solutions for the life science market including sample management, small volume liquid handling and detection systems.

Biography:

Emi Hifumi has obtained her PhD from Kyushu University and studied Antibody Engineering and Catalytic Antibody. She was a Research Assistant at Hiroshima Prefectural University and is currently a Professor at Oita University from 2007. She has published more than 30 papers in reputed journals.

Abstract:

We are developing catalytic antibody light chains (human antigenases) by using the human genes belonging to subgroup II, which exhibit some unique features such as enzymatic function and also anti-virus infection. We amplified and cloned cDNAs encoding the human antibody light chains (kappa) belonging to subgroup II. The obtained cDNAs were transformed into E. coli and then expressed as the protein. The highly purified (over 95%) antigenases were submitted to the following experiments. Several antigenases out of over 200 antigenases investigated showed the suppressive effect on the infectivity of influenza virus H1N1 not only in vitro but also in vivo assay. 22F6 antigenase clearly prevented from the infection of influenza virus H1N1 in vivo, where PR-8 strain was used. The serum titer of the mice inoculated with antigenase treated virus was substantially low even at 21 dpi, comparing with the positive control, suggesting the lost of antigenicity of the virus. Taking into account that the antigenase showed the catalytic activity as DNase and RNase, the loss of the infectivity may be due to the cleavage of the virus RNA by the antigenase. On the other hand, 23D4m also possessed a suppressive function for the infection of influenza virus in vivo, while it is a monomeric light chain. In the investigation of nasal inoculation schedule of 23D4m, it clearly showed anti-viral effect under the simultaneous inoculation of the virus and the light chain. These results suggest that the above antigenases have the high possibility to prevent from the infection of influenza virus.

Biography:

Enrica Pessione has completed her PhD in Microbiology at “Institut Pasteur”, Paris. She has been a Researcher in Microbial Biochemistry at Turin University getting expertise in enzymology, metabolic biochemistry and microbial proteomics. Since 2004, she is an Associate Professor in Biochemistry and Member of the Italian Microbial Biotechnologies Association. She has focused her interest on metabolic pathways important for food safety control or to establish the probiotic potential of lactic acid bacteria. She has published over 80 scientific articles. She is in the Editorial Board of Journal of Integrated Omics.

Abstract:

Lactic acid bacteria (LAB) have extensively been used as well-defined starters in the industry of fermented food production. More recently, their role as biocontrol agents to counteract food-borne infections and spoilage bacteria has also been proposed, to lower the amount of salts, sugar and preservatives in food, but also to reduce the cold-chain need, with benefits for costs and sustainability. Nowadays, LAB potential as probiotics and nutraceutical vectors has a huge impact on the pharmaceutical and food supplement industry. Although it is very promising to use LAB as microbial cell factories for functional food production, however, their metabolic profiles are often underexplored and most products underexploited. Proteomics is a promising tool for obtaining information on the metabolic pathways of interest and how these routes can be modulated by exogenous conditions. This presentation will refer all the cutting-edge products that can be obtained by LAB, the use of comparative proteomics and sub-proteome techniques to ascertain probiotic safety and efficacy, nutraceutical compounds release and optimization of the industrial production. In particular, selenium fixing ability, neuroactive compounds production, food-encrypted peptides release, antimicrobial molecules and exopolysaccharide (EPS) biosynthesis as well as bacterial resistance technological treatments will be considered.

Biography:

Mohit Kapoor is a Senior Scientist at the Krembil Research Institute, University Health Network in Canada. He leads the Cartilage Biology and Joint Restorative Medicine Research within the Arthritis Program at the University Health Network. He has published over 60 research article, reviews and book chapters in high impact journals. He is a recipient of several prestigious Research and Fellowship Awards from various research organizations across the globe. He is also an Editor of book on osteoarthritis that was recently published in 2015 and sits on Review Panel of various research funding organizations.

Abstract:

Osteoarthritis (OA) is the most common form of arthritis and ageing-related joint pathology associated with degradation of the articular cartilage, synovial inflammation/fibrosis, subchondral bone remodeling and osteophyte formation. This disease results from alterations in the joint tissues that lead to pain, loss of motion and instability. The economic burden associated with this disease is substantial. Specific mechanisms associated with the joint destruction and associated pain during OA is largely unknown. Due to the lack of biomarkers, it is impossible to identify patients exhibiting early stages of OA, leading to severe joint destruction. Furthermore, due to poor understanding of the underlying disease mechanisms, no disease-modifying therapies to treat OA exist. MicroRNAs (miRNAs) are small non-coding RNAs that are expressed as primary stem loop precursors and undergo maturation by enzymatic processes. It is estimated that miRNAs regulate more than 60% of all coding genes and play pivotal roles in pathophysiological processes; including cell proliferation, differentiation, genomic stability, metabolism, apoptosis and aging. In this lecture, I will talk about how we employed a combination of gene expression analyses, computational biology and in vitro biological functional assays to identify panel of microRNAs as potential biomarkers and therapeutic targets in osteoarthritis.

Biography:

Arata Katayama has completed his PhD from Tokyo Institute of Technology and worked as Postdoctoral fellow at University of California, Institute of Toxic Substances Research. Presently, he teaches Sanitary Engineering for undergraduate course and Ecotoxicology for graduate course as a Professor of Department of Civil and Environmental Engineering, Nagoya University, Japan. He also serves many governmental committees on environmental pollution, polychlorinated biphenyl remediation and monitoring of environmental quality.

Abstract:

Microbial remediation is one of the promising technologies for treating contaminated environment, especially anaerobic technology for the aquifer and sediments. Anaerobic microbial remediation does not require the soil excavation and/or aeration of aquifer, therefore, it is less expensive and energy saving. However, microorganisms tend to have a narrow range of degradation spectrum of toxic substances, although the polluted sites often contain multiple toxic substances. To this shortcoming of microbial capacity of detoxification of multiple toxic substances, we have conducted the study on a synthetic microbial community to widen the microbial spectrum in the degradation of toxic substances. In this study, we have combined the halogenated aromatics-respiring anaerobe (Dehalobacter), halogenated aliphatics-respiring anaerobe (Geobacter), aromactics-oxidizing anaerobe (Desulfatiglans) and hydrogen-producing anaerobe (Clostridium). The synthetic community was also conducted using multiple enriched cultures including these anaerobes. The experiments suggested that the combination of different microbial strains/community is successfully carried out to widen the degrading spectrum. Toxic metabolites from one anaerobe, hydrogen sulfide from sulfate-reducing anaerobe, caused the inhibitory effect on other members in synthetic anaerobe community. In the case of synthetic anaerobic community applied to the plug-flow reactor, pentachlorophenol, the examined pollutant was completely mineralized. All the microbial strains distributed into the narrow upstream area in the reactor based on the detection of functional genes. These results suggested that the design and construction of synthetic anaerobic community would be useful and important for the successful bioremediation.

Biography:

Sharon Mendel Williams joined Coventry University as a Lecturer in the School of Life Sciences in November 2014. She worked as a Post-doctorate Research Fellow in both Chemistry and Biology departments of Warwick University for 8 years. Her research focuses on the biophysics and biochemistry of proteins, and understanding the mechanisms of enzymes. She has a wide range of depth and experience in molecular biology, biochemistry, and chemistry. She is a member of the Royal Society of Chemistry and has been awarded a grant from the RSC research fund to accomplish the current research.

Abstract:

Lignin is an organic polymer found in the cell walls of plants. Lignin can be used to create biofuels, or as an organic hydrocarbon source for a large variety of chemicals and polymers. However, lignin is very robust and current industrial processes for using it are inefficient. Therefore, a useable biological process for degrading lignin would be of great benefit. Understanding the pathway of lignin degradation, and the byproducts, is essential in order to be able to exploit the use of micro-organisms. Furthermore, we can characterize novel bio-products obtained by enzymatic oxidation of lignin, which could have very interesting applications for industrial biotechnology. In the current project Dyp-type peroxidases from Gram-negative Pseudomonas fluorescens Pf-5 and recombinant Sphingobacterium MnSOD1 and MnSOD2 cloned into E. coli and were investigated. These are bacterial enzymes that are already known to degrade lignin. Different genetic mutations were introduced, and the resulting enzymes were characterized by using them with different lignin substrates. The reaction compounds were analysed by reverse phase HPLC/ GC-MS. The goal is to improve the effectiveness of the enzymes, increase the production of the enzymesand degrade the lignin into different and more useable compounds. Any of these goals would be of valuable scientific and commercial benefit.

Biography:

Betty Lee has a PhD from Dartmouth Medical School, USA, MS in Clinical Chemistry from the University of Windsor, Canada and MS in Biochemistry from LSU Medical Center, USA. She completed her Post-doctoral training at the National Institutes of Health, USA. She currently works as a Licensing Officer with the US Government. She educates industries and academia about the Export Administration Regulations (EAR) and participates in outreach. In addition, she participated in the policy review of the Executive Order titled “Optimizing the Security of Biological Select Agents and Toxins in the United States” signed by President Obama on July 2, 2010.

Abstract:

Balancing biosecurity and legitimate life science research has become a priority in recent years. The rise of biotechnology and informatics has made rapid advances in the 21^st century. Such a convergence of biology and technology increases the pace of biological findings and the emergence of new technologies. Emerging technologies that could be used for biological warfare poses a formidable challenge because of the unpredictable nature of science. Dual use research in the life sciences requires some oversight by the government and funding agencies. The published results of scientific research could be used to improve health or agricultural products or they could be used to enable bioterrorism. Life Sciences research is conducted increasingly in an interdisciplinary and international environment. Informatics, systems biology, nanotechnology, and synthetic biology are at the forefront of such endeavors. Oversight of such research is essential to continue the free exchange of information and also balance the national security concerns. One of the tools that is at the nexus of biosecurity and life sciences is export control. The session will provide an overview of the biological agents (viruses, bacteria and toxins), genetic elements (DNA, plasmids, vectors) which are currently controlled on the Australia Group Control List and Commerce Control List for exports. The Australia Group is an informal forum of countries whichseeks to ensure that exports do not contribute to the development of chemical or biological weapons. Topics of discussion will include deemed exports, fundamental research and technology transfer.

Biography:

Anthony C Forster has discovered the hammerhead catalytic RNA structure, invented external guide sequences for ribonuclease P and created unnatural genetic codes de novo, all of which founded biotech companies. He has published in journals including Cell, Nature and Science, edited volumes of Methods and Biotechnology Journal and coauthored "Synthetic Biology: A Lab Manual."

Abstract:

We have developed simplified, purified, bacterial translation systems to facilitate studies of substrate recognition in protein synthesis and enable new applications. Surprises in translation include slower incorporation of proline and other N-alkyl amino acids and that peptide bond formation, not accommodation, is rate-limiting in dipeptide synthesis. One application is directed evolution in vitro of small-molecule, peptidomimetic drug candidates by redesigning the genetic code for the synthesis and display of polymers containing unnatural amino acids. Fast kinetics with unnatural amino acids has identified the causes of several inefficiencies and lead to ways of improving incorporation yields. Another application is cost-effective, scalable, purified, in vitro translation. The latter was achieved by total synthesis and BioBrick assembly of a 58-kbp module encoding 30 translation factor cistrons, breaking new ground in de novo design for synthetic genomics.

Biography:

Martin Falk has completed his PhD from Masaryk University in Brno, CR. He is the Leader of the Department of Cell Biology and Radiobiology at the Institute of Biophysics of the Czech Academy of Sciences. He has participated in more than 30 papers that concern the role of chromatin structure in regulation of cellular processes. His research interests include DNA damage and repair, carcinogenesis, tumor cells radio-sensitization and radiobiology.

Abstract:

Until recently, mainly the genetic and biochemical aspects of processes in the cell nucleus were studied. Nowadays, in the advanced era of 'omics', we have already obtained substantial information on how dozens of proteins interact in the frame of complex cellular signaling pathways and networks. However, additional levels of complexity, like chromatin spatio-temporal organization, have recently emerged as important regulators of fundamental nuclear processes. In this lecture, we will focus on the maintenance of genome integrity, namely the repair of DNA double strand breaks (DSBs). The integrity of the human genome is continuously threatened by intercellular and environmental factors and DSBs represent the most serious DNA lesions; even a single DSB can initiate cell death or cancer when repaired inaccurately. On the other hand, tumor cells are most efficiently killed by DSBs introduced by radiotherapy or chemotherapy. In the last decade, we studied how DSBs, caused by different radiations (high-LET, low-LET) are being induced and repaired. We will discuss the complexity of DSB repair with emphasis on the question of how chromatin structure (nuclear architecture) influences its mechanism and fidelity. Based on the results obtained, we propose a model of the relationship between the higher-order chromatin structure, DSB induction and repair and formation of (carcinogenic) chromosomal translocations.

Biography:

Nobuo Fukuda has completed his PhD from Kobe University. He is a Senior Research Scientist of Biomedical Research Institute, AIST. He has published more than 15 papers in reputed journals.

Abstract:

Sake yeasts belong to the budding yeast species Saccharomyces cerevisiae and have high fermentation activity and ethanol production. Although the traditional crossbreeding of sake yeasts is a time-consuming and inefficient process due to the low sporulation rates and spore viability of these strains, considerable effort has been devoted to the development of hybrid strains with superior brewing characteristics. In the current study, we constructed growth selection systems for ‘a’ and α-type cells derived from parental MATa/α yeasts and confirmed that the generated cells possess suitable mating abilities for the production of hybrid yeasts. To achieve it, we designed suitable genetic circuits for expression of the kanMX4 selection marker gene to permit isolation of ‘a’ and α-type cells, respectively, on solid medium. And also, we prevented autopolyploidization of yeast cells derived from the same parent via forced expression of the a1 or α2 gene to increase hybridization efficiency in crossbreeding. Industrially-used strains Kyokai No. 6 and No. 7 were selected as parental sake yeasts and we generated a hybrid strain, designated K76, using the constructed selection systems, which inherited the genetic characteristics of both parents. Notably, because all of the genetic modifications of the yeast cells were introduced using plasmids, these traits can be easily removed. The approach described here has the potential to markedly accelerate the crossbreeding of industrial yeast strains with desirable properties.

Biography:

James M Carothers is currently an Assistant Professor at the University of Washington and an Investigator of the Engineering Biology Research Consortium. Previously, he was a Postdoctoral Fellow with pioneering synthetic biologist Jay D. Keasling at the University of California Berkeley. He has earned his PhD at Harvard University with 2009 Nobel Prize-winner Jack W Szostak. He has a BS in Molecular Biophysics and Biochemistry from Yale. His co-authored papers have been cited more than 1300 times and his recent work in synthetic biology has been recognized by the University of Washington Presidential Innovation Award and the Alfred P. Sloan Research Fellowship.

Abstract:

Programmable RNA devices and systems can be engineered for applications in biosensing, information processing and dynamic genetic control. In principle, by combining advanced computational simulations with massively-scaled experimental analysis we investigate the limits of RNA device and system design and engineer synthetic metabolisms to produce medically and industrially-important materials. Here, I will present results using new designable molecular architectures for engineering ultrasensitive RNA aptamer nanosensors as technologies for parallelized metabolic output profiling, a novel class of riboswitch-aptazymes as dynamic metabolite-responsive feedback controllers and programmable guide RNA (gRNA) expression platforms for implementing complex CRISPR-dCas9-based transcriptional networks.

Biography:

Mark S Friddin has completed his PhD from the University of Southampton. He has joined the Ces group at Imperial College London as a Postdoctoral Research Associate working on the CAPITALS program in January 2015. His research interests focus on the microfluidic assembly of model membranes, the characterization of ion channels using electrophysiology and the development of smart synthetic microsystems for drug discovery.

Abstract:

Droplet interface bilayer (DIB) networks are becoming increasingly considered as powerful minimal-tissue constructs, however a bottleneck preventing the advancement of this technology is the difficulty of positioning cell-sized droplets into customizable 3D architectures. We address this problem by showing the use of optical tweezers to precisely assemble individual microdroplets ≤20 µm in diameter into complex user-defined 2D and 3D DIB networks. We achieve this by adding sucrose to the aqueous phase to reverse the refractive index contrast of the water in oil microdroplets. This allows us to directly trap the microdroplets in 3D as opposed to exploiting fluid motion generated by the thermocapillary effect or the Marangoni convection effect as reported previously. DIB connectivity between the optically manipulated droplets was confirmed by demonstrating the interdroplet exchange of calcium ions through alpha hemolysin (αHL) nanopores inserted into the membrane. To showcase our ability to method to assemble single and multilayered DIB networks, we constructed both linear and branched symmetric/asymmetric 2D networks together with a 3D droplet tower composed of 11 cell-sized droplets. To our knowledge, the DIB networks assembled using our technique is among the smallest and most complicated reported to date. We envisage that this technology will pave the way for the development of a new generation of minimal-tissues, smart drug delivery systems and bio-electronic circuits assembled from modular droplet components.

Biography:

Hector Hugo Caicedo holds PhD in Bioengineering from the University of Illinois at Chicago. He is the recipient of MIT, Bogazicy University, Antalya University and Universite Pierre and Marie Cuire Pre-doctoral Fellowships as well as Harvard-MIT/HST Post-doctoral Fellowship. He has several publications including several peer-reviewed papers, one book chapter and a provisional patent application. Additionally, he has been awarded more than 20 recognition awards including three National Science Foundation Fellowships in the US and the Mayor’s Civic Merit Medal of Cali given directly by the President of Colombia. Currently, he is a Scientist and Scholar at Janssen R&D, Pharmaceutical Companies of Johnson & Johnson. His interdisciplinary scientific research interests range from material science and bioengineering to cell biology, medicine and pharmaceutical research. At J&J, he is a Subject Matter Expert in Microfluidics Science & Technology and conducts research on drug lead identification, characterization and optimization. Additionally, he is also a Scholar Trainee at the Corporate Sustainability and Innovation program at Harvard University and serves as the Director of Research and Outreach at the US National Biotechnology and Pharmaceutical Association.

Abstract:

Microfluidics has the potential to influence all subject areas of biotechnology from agricultural and environmental to medical and pharmaceutical biotechnology. Even though translational microfluidics science and technology are still at an early stage of development, the pharmaceutical biotech industry may represent the best sector for microfluidics to thrive. With the advent of single-cell analysis, immunotherapy, genetic engineering, micro-scale immunoassays, precision medicine and disease interception; innovative lab-on-a-chip solutions are shaping biotechnology research in ways that are not possible using traditional methods. These solutions enable scientists to shed light on the mechanisms that regulate a myriad of highly intricate biological processes, in both normal and disease states. Here, we highlight some microfluidics-based science and technology that we have developed to enable cutting edge biomedical research, including microfluidics-based real-time tracking of neuronal protein in isolated axons, SAW technology and method development to support pharmacokinetic and pharmacodynamics studies. We also challenge the current paradigm that is focused on searching for a “killer application” as a response for the poor adoption and commercialization of microfluidics technology beyond academic engineering laboratories. Based on our own scientific experience, both in academic and industry laboratories, we provide several reasons that might explain the poor adoption of microfluidics in mainstream biomedical research and the biotech industry. Additionally, we suggest prospects for future directions that could help address some of the present challenges. The aim is enhanced translational microfluidics research that tackles real-world applications in the life sciences and pharmaceutical biotech industry to help develop innovative biotherapeutics for human healthcare.

Biography:

Krishnan is a Senior Lecturer in Chemical Engineering and in the Centre for Process Systems Engineering and affiliated with the Institute of Systems and Synthetic Biology at Imperial College. He has completed his undergraduate studies at IIT-Madras, PhD at Princeton University (both in chemical engineering) and was an Associate Research Scientist in Electrical Engineering at the Johns Hopkins University.

Abstract:

Biochemical modification pathways and cascades are the building blocks of signal transduction and are usually studied, analyzed and understood in temporal (lumped) terms. However it is becoming amply clear that in many cellular contexts, reactions occur at different locations and that the spatial aspects of signal transduction and biochemical pathways in general are especially important. This aspect is however poorly understood for multiple reasons. In this talk we focus on the effect of compartmentalization, which is ubiquitous in cellular systems. The effects of compartmentalization are important in understanding concrete pathways and cellular processes which involve compartmentalization, how this has been employed by evolution and also for designing microcompartments in synthetic biology. In this talk, I will discuss the effects of compartmentalization in a range of biochemical modification cascades/pathways: (1) Enzymatic cascades where the output at one stage is an enzyme for the next stage, (2) Pathways where the output at one stage is the substrate for the next stage, (3) Phosphorelays and (4) Open pathways. In each case, a systematic analysis of the effects of compartmentalization is made by comparing the spatially distributed system (modeled by PDEs) with the colocalized system (modeled by ODEs). Through this systematic analysis, we uncover a whole range of effects of compartmentalization of such pathways. We then discuss the relevance and implications of these results for engineering spatial compartmentalization.

Biography:

Pauli Kallio has completed his PhD at the Department of Biochemistry at University of T urku, Finland (2008), studying the secondary metabolic pathways and enzyme-level determinants behind the diversity of bioactive compounds in   Streptomyces  bacteria. Currently he leads a  Synthetic Biology  Group in the Plant Molecular Biology Unit at the University of Turku, funded by the F innish Funding Agency for Innovation (Tekes), with a focus on biofuel research and synthetic biology applications in photosynthetic cyanobacteria. His primary fields of expertise include  molecular biology , in vitro enzymology and bacterial metabolic engineering.

Abstract:

     Cyanobacteria      are a diverse group of ph otosynthetic     prokaryotes     which have b een extensively studied for their potential as advanced     biotechnological     hosts for the production of different    chemical    compounds, namely biofuels a  nd their precursors. The ultimate advantage of    cyanobacterial    production platforms would be the capacity to generate desired end-products directly from atmospheric CO₂ using sunlight as the sole energy without the need for externally supplied sugar as a substrate. Despite the pre-eminent advantages, the use of such production    systems    is still limited by inefficiency and low production yields, wh ich render the applications commercially non-competitive. To overcome these barriers, our current research is focused on the development and evaluation of a versatile   molecular biology   toolbox using  synthetic biology  approaches, in order to effectively harness the photosynthetic capacity to our advantage. Collectively, we aim at developing a palette   of standardized validated tools for the assembly and optimization of complex  multi-gene  over-expression systems in cyanobacteria, which we currently are missing. This will allow us to find, characterize and compare t he most suitable genetic elements and expression strategies for our needs and consequently, to exploit our extensive fundamental understanding on the photosynthetic machinery to re-route the metabolic flux towards the target meta bolites more efficiently. In our view, systematic synthetic biology approaches combined with the biosynthetic potential of photosynthetic microorganisms may contribute to the development towards sustainable bioeconomy independent of petroleum-based fuels.

Biography:

Sasa Rezelj has completed her MSc from   Biotechnical   Faculty of the University of Ljubljana. Sh e is the currently a PhD student in Department for  Molecular Biology  and Nanobiotechnology, National Institute of Chemi  stry, Slovenia. Her area of interest is synthetic biology approach to study and engineer listeriolysin O under supervision of Professor Dr. Gregor Andreluh. 

Abstract:

     Listeriolysin O      (LLO) is the pore forming toxin and the most important virulence factor of bacteria      Listeria monocytogenes     , which causes food-borne disease listeriosis. It belongs to cholesterol-dependent cytolysins (CDC), a family of pore-forming toxins produced primarily in Gram positive bacteria. LLO enables      bacteria      to escape from host     phagolysosome     and to spread into the neighboring cells. Moreover, LLO is a unique protein among CDC, since its stabil ity is pH dependent. Pore forming    proteins   , such as   α-hemolysin  , have shown   their way to biotec  hnological applications (e.g., DNA sequencing through nanopore). LLO also found its potential use in medicine as a vaccine adjuvant and carrier   molecule   for anti-tumor therapies and  gene  delivery. We believe that by revealing the exact mechanism of LLO pore forma tion, we can engineer LLO to form pores, which can be manipulated and use d  in biotechnological applications. First, we used giant  unilamellar vesicles  (GUVs) to study LLO’s mechanism of pore formation. Confocal fluorescence microscopy and flow cytometry were used to analyze time, concentration and environmen t dependent pore formation of LLO. We discovered the damaging effect of LLO’s pore formation on membranes by analyzing the permeability of GUVs for fluorescent dextrans (FDs) of different sizes and populations of GUVs upon LLO addition. LLO pore forming activity enabled time dependent permeabilization of FDs up to 150 kDa, which is larger than previously assumed pore diameter. LLO was able to damage the vesicles in such extend, that it led to their destruction. Furthermore, we used cell free in vitro synthetic biology approach to construct and produce LLO inside synthetic phospholipid vesicles, which can be specifically controlled by environmental changes.

Biography:

Reem Swidah has completed her PhD from Manchester University in 2016 and she has been working with her supervisors Mark Ashe and Chris Grant on biofuel production from yeast. She has published her first paper in Biotechnology for Biofuels.

Abstract:

Biobutanol represents a second generation   biofuel  , which can be produced naturally by a number of   microorganisms  . This alcohol has a number of significant advantages over bioethanol in terms of its physical properties as a fuel, but production   systems   suffer from various drawbacks. Therefore, we sought to transplant an entire butanol production pathway (the ABE pathway) into a   Saccharomyces cerevisiae   strain. However, this pathway was incapable of generating reasonable yields of butanol without further metabolic alteration to channel carbon towards the substrate of butanol production,   acetyl CoA  . For instance, the major alcohol dehydrogenase, ADH1, was deleted and two enzymes involved in acetyl-CoA   biosynthesis   were overexpressed to give strains capable of producing 300 mg/L butanol. Surprisingly, deletion of the ADH1 gene alone is sufficient to produce 40 mg/L butanol from an endogenous pathway. Previously, this endogenous butanol production pathway was characterized and proposed to derive from the    mitochondrial   catabolism  of threonine via multiple leucine  biosynthetic genes  and the conversion of 2-ketovalerate to butanol. Therefore, in the studies described here, we aimed to understand the relative contribution of exogenous and the endogenous pathways of butanol  synthe sis  to overall butanol yields. Work will be presented to suggest that both pathways are active in yeast  adh1D mutants , but that the endogenous route for butanol synthesis is weaker and does not use the pathway previously proposed via  Leucine  metabolic enzymes . This work therefore makes use of synthetic biology and metabolic engineering to effectively set the scene for an initiative toward s higher yields of butanol in yeast via concerted interventions in both the endogenous and exogenous pathways.

Biography:

Chiara Gandini has completed her MSc in Industrial    Biotechnology    in 2012. She  is Coauthor of an international patent. She has been working in   Biochemistry   Laboratory of Department of  Life Sciences  and Systems Biology, U   niversity of Torino, on microorganism’s metabolic engineering since 2012. 

Abstract:

   Consolidated Bioprocessing    (CBP) is a key feature of the so-called 3^rd generation    biorefinery    consisting of cost-sustainable single-step (d irect) fermentation of   cellulosic biomass   into industrially rele vant products such as biofuels (notably, ethanol and or butanol). No natural   microorganism   isolated so far can produce  biofuels  directly from cellulose with the efficiency required by industrial processes.    Clostridium cellulovorans  is an anaerobic bacterium among the most efficient plant biomass biodegraders. It efficiently hydrolyzes cellulose, xylan and pectin, but its main catabolites are  organic acids , while little ethanol is biosynthesized. The final aim of the present study is the development of recombinant ethanol and or butanol hyper producing  C. cellulovorans strains by metabolic engineering. Achievement of this purpose is currently hampered by major hurdles. Nowadays, no detailed study on C. cellulovorans central metabolic pathways exists. Furthermore, no specific tools for C. cellulovorans chromosomal gene knock out/in are available. The present contribution was focused on the development of a model of the C. cellulovorans central metabolism by integration of metabolomics, transcriptomics and proteomics data. Attention was mainly given to comparison of C. cellulovorans metabolic network during growth on cellulose with respect to cells growing on soluble sugars. As far as the development of optimized gene tools for C. cellulovorans is concerned, different transformation protocols were compared. Furthermore, the ClosTron methodology was used as a basis for developing reliable protocol for gene integration in the C. cellulovorans chromosome. These results will be a key for future C. cellulovorans metabolic pathway engineering aimed at direct conversion of cellulose to biofuels.

Biography:

Loredana Tarraran has obtained her MSc in Molecular Biology from Turin University. She is interested in environmental field and working on metabolic engineering of microorganisms for application in biorefinery since 2009. She is author of one national congress communication and Co-Author of other three national and three international congress communications. Furthermore she is Co-Inventor of one international patent.

Abstract:

      Lactic acid       (LA) is an extensively employed chemical. Notably, one of its main current applicati ons is for      synthesizing      biocompati ble and biodegradable plastic biopolymers, i.e.,      polylactide      (PLA), poly (lactic acid-co-lysine), poly (lactic acid-co-glycolic acid). The latter can be used for clinical application (i.e.,  tissue engineering) and especially PLA as packaging thus replacing traditional plastics. Lactic acid bacteria are the main natural LA producers. Both “green” property and cost-sustainability of LA-based polymers can be improved if LA is obtained by fermentation of abundant      cellulose     -based wastes produced by our society, such as municipal solid waste and agricultural by-products. In this light, the development of a     microorganism     able to both use    cellulose    as fermentation substrate and produce LA can achieve both environmentally an d economically sustainable   biopolymer   production. The aim of thi s work  was to obtain a recombinant     Lactococcus lactis    able to depolymerize cellulose by expressing heterologous    proteins    derived from the   cellulolytic   complex, i.e., the  cellulosome  of    Clostridium cellulovorans .  C. cellulovorans  cellulosome consists of enzyme subunits attached to a non-catalytic scaffold  protein . Such scaffoldin is anchored to  C. cellulovorans cell surface and allows optimal overall enzyme spatial organization, close to both cellulosic substrate and cell surface thus leading to improved cellular intake of cellulose hydrolysis products. In this work four different recombinant scaffoldins were constructed by molecular assembling of different protein domains of C. cellulovorans cellulosome. Engineered scaffoldins were successfully expressed by L. lactis. Their presence in cytosolic, extracellular and cell-wall fractions of L. lactis and their interaction with cellulosomal enzymes were analyzed by multiple approaches.

Biography:

Denis Kalemasi has completed his MSc in Industrial Biotechnology at the University of Turin in 2015. He has been working since 2014 on the construction of a recombinant L. lactis for the direct conversion of cellulose into L-lactic acid.

Abstract:

One of the possible applications of   synthetic biology   is the “creation” of recombinant bacteria able of producing building blocks. The latter have great potential since they can be used for the production of bio-bas ed polymers possibly replacing oil-derived plastics.  Lactic acid  (LA) is among the most requested compounds by the ch emical industry and is already largely employed for the production of its  polyesters  (i.e., polylactides), which are general purpose biodegradable and biocompatible plastic materials. Most LA produced worldwide is obtained by lact ic acid bacteria (LAB)-based fermentation. However, current processes depend on expensive feedstocks (e.g., glucose) or compete for food crops (e.g., corn). Cellulose, instead, is an ine xpensive substrate found in plant material-derived waste. Our aim is to construct a strain of Lactococcus lactis able to produce LA directly from cellulose (that is without prior enzymatic saccharification by commercial cellulases) by metabolic engineering. Only relatively few natural bacteria are able to grow with only cellulose as the substrate. Clostridium cellulovorans is one of them. To achieve our goal, we’ve introduced components of C. cellulovorans cellulose depolymerizing machinery into Lactococcus lactis. In particular, in this work we have shown that, by introducing only two C. cellulovorans glycosyl hydrolases, we were able to develop a recombinant L. lactis able of metabolizing short chains of cellulose (i.e., cellooligosaccharides with degree of polymerization up to 10) into L-LA with a yield close to 100%.

Biography:

Dr. Ingy Moustafa Hashad’s work involved the study of “contribution of the p22 phox gene of NAD(P)H oxidase and eNOS gene polymorphisms in the predisposition of early onset acute myocardial infarction in Egyptian population”. She learnt several techniques through her research including, conventional PCR and SNP analysis, ELISA, electrophoresis, and spectrophotometric determinations. She finished successfully her Ph.D. in May 2012 with great appraisal from the supervisors and examiners. Later, she had the opportunity to spend three months summer research in the research lab of Prof. Wolfgang Poller, Charité Centrum für Herz-, Kreislauf- und Gefäßmedizin and another three months in the research lab of Prof. Michael Bader, Max-Delbruck Center for Molecular Medicine, Berlin, Germany which added to her scientific and technical knowledge and skills.

Abstract:

Background:    Cardiovascular diseases    (CVD) are the universal cause of morbidity and the leading contributor to mortality in both developed and developing countries nowadays. Connexin (Cx)    proteins    are the building blocks of gap junctions.  Among these, Cx37 and Cx40 have been found to be expressed on vascular system and reported to have a cardio protective role.   Glutathione peroxidase-1   (GPx-1) enzyme and  Manganese Superoxide Dismutase  (Mn-SOD), represent a defense mechanism against oxidative stress, thereby c ontributing to the prevention of atherosclerosis. Besides, high  homocysteine  levels (Hcy) and  Hexanoyl Lysine adduct  (HEL) are considered to be an independent risk factor for wide range of diseases such as CVD and their complications including Acute    Myocardial Infarction    (AMI). AMI is inflammatory pathology, including cytokines such as Fractalkine which plays a central role in   inflammation   and tissue injury.  

Subjects & Methods: A total of 205 Egyptian subjects were recruited for the study. They were divided into 105 AMI patients and 100 healthy controls.   Genotypes   for each participant were determined using a  polymerase  chain reaction restriction fragment length polymorphism (PCR-RFLP) for the Cx37, Cx40, GPX-1 and Mn-SOD genes.  Serum levels of sVCAM-1, HE L adduct, Homocysteine and fractalkine were detected quantitatively using ELISA.

Results: Allele frequencies for both Cx37 and Cx40 were not significantly different between AMI patients and controls (p=0.93 and 0.24, respectively). The  genotype  distribution for GPx-1 gene was not significantly different between the AMI patients (CC 56.7%; CT 41.7%; TT 1.7%) and control subjects (CC 53%; CT 45%; TT 2%), (P=0.6008). The  prevalence of the V/V genotype of the Val16Ala of Mn-SOD gene polymorphism was significantly more frequent in AMI patients than in control subjects (p=0.0142). In addition, Serum levels of sVCAM-1, HEL adduct, Homocysteine and fractalkine were significantly elevated in AMI patients compared to control subjects (p=0.0273).

Conclusion: Contribution of inflammation and oxidative stress markers in addition to Mn-SOD gene polymorphism in the pathogenesis of AMI in Egyptian Population.

Biography:

Azizah Nahari is a PhD student in her last year from the University of Edinburgh. She has a Master’s degree from King Abdulaziz University. She is working as a Lecturer at King Abdulaziz University School of Biological Science.

Abstract:

Limitation of crop productivity by     phosphate     (Pi) is widespread and will probably increa se in the future. A better understanding of    phosphate    use efficiency (PUE) is required for    engine ering    nutrient-efficient. In this study, we design ed a   hydroponic   syst em for a specific reason which is to avoid complication from differences in Pi uptake. The current experiment examined 2 13 of the total 527    MAGIC     lines under high and low   phosphate   conditions in hydroponic  system, and the results generated considerable variance with regard to various PU E elements and growth features (namely, RFW, SDW, SPC, and flowering time). By using   QTL mapping  , we have identified QTLs involved in PUE. Having examined the broad 213  MAGIC  popula  tion, considerable variance in PUE and its constituent features has been recorded. This is indicative of the manifest variance in  PUE  for  the examined lines. It is significant to note that, at the level of the species, lines under the low Pi condition displayed greater PPUE when considered in relation to those under the high condition. Fur  thermore, the association with regard to  PPUE  and the features of SDW and Pi content was determined as positive fo r all lines under the low Pi condition. Notably, these findings provide valuable insight regarding individual lines and the degree to which they can grow effectively in high and low Pi conditions. Additionally, it has identified lines characterised by dissimilar PUE values that could be appropriate for utilisation in breeding schemes with the intention of enhancing the features in novel cultivars.

Biography:

Anna K Stavrinides has completed MSc in Plant  Genomics  and Crop Improvement and she is currently undertaking PhD in the Lab of Prof Sarah O’Connor both at University of East Anglia a nd the John Innes Centre. She has Bachelors degree in Molecular Biology from the Université Montpellier 2. Her interests are  focused around plant abiotic stress resistance and plant/environment interactions. 

Abstract:

Monoterpene indole  alkaloids  are a diverse family of compounds, some of which are of economic importance as drugs for use in the clinic. The medicinal plant  Catharanthus roseus  (Madagascar periwinkle) produces more than 100 monotepene indole alkaloids and is the source for two  anti-cancer  alkaloids vinblastine and vincristine. Biosynthesis of these alkloids is long and involves >20 enzymatic steps and multiple tissues and cell compartments, which hav e complicated the discovery of the biosynthetic enzymes necessary for production of these useful alkaloids. The central intermediate for generation of the various monoterpene indole alkaloid scaffolds is strictosidine, which can be found in many plants, primarily of the Apocynaceae and Gelsemiaceae families. This alkaloid is deglycosylated and the resulting compound is reactive and unstable. Monoterpene indole alkaloids with various backbones can be obtained from this precursor and are found in different species. Recently we discovered the first enzyme which can take this reactive precursor and reduce it to form a heteroyohimbine backbone (Tetrahydroalstonine synthase). This discovery completes the biosynthesis of an alkaloid found in many plant species which produce strictosidine such as Rauvolfia serpentina. I will present our recent work on the elucidation of this dynamic branch of the monoterpene indole alkaloid pathway. Using the information we have gained about the system (gene expression, protein interaction, localization, enzyme specificity) we propose an integrated hypothesis for the evolution of the heteroyohimbine biosynthetic enzymes in the medicinal plant Catharanthus roseus.