Difference between revisions of "Team:Virginia/Contribution"

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             <div>General Template Page</div>
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             <div>Contributions</div>
 
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           <div class="sectionTitle" id="Section 1">Section 1</div>
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           <div class="sectionTitle" id="Section 1">iGEM Resource Page</div>
 
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                 The lack of a versatile and reliable way to improve metabolic flux channeling, pathway orthogonality, and product yields is a major impediment to the expanded utilization of biosynthesis for the production of drugs and industrially valuable chemicals. Manifold, a platform technology that addresses this problem, consists of <div class="dict">bacterial microcompartments<span><img src="https://upload.wikimedia.org/wikipedia/commons/thumb/2/25/Carboxysome_and_bacterial_microcompartments.jpg/800px-Carboxysome_and_bacterial_microcompartments.jpg"/>Bacterial microcompartments (BMCs) are organelle-like structures, consisting of a protein shell that encloses enzymes and other proteins. BMCs are typically about 40–200 nanometers in diameter and are entirely made of proteins. The shell functions like a membrane, as it is selectively permeable.</span></div> (BMCs) with encapsulated dsDNA scaffolds <div class="ref">[1]<span>Elbaz, J., Yin, P., &amp; Voigt, C. A. (2016). Genetic encoding of DNA nanostructures and their self-assembly in living bacteria. Nature communications, 7(1), 1-11.</span></div> that sequester and spatially organize, at fixed concentrations, biosynthetic enzymes presented as zinc-finger fusion proteins. Here we deliver the designs for an E. coli cell capable of synthesizing resveratrol using the Manifold platform. The Manifold platform will help lower costs and expand the applications of chemical biosynthesis. The lack of a versatile and reliable way to improve metabolic flux channeling, pathway orthogonality, and product yields is a major impediment to the expanded utilization of biosynthesis for the production of drugs and industrially valuable chemicals. Manifold, a platform technology that addresses this problem, consists of bacterial microcompartments (BMCs) with encapsulated dsDNA scaffolds that sequester and spatially organize, at fixed concentrations, biosynthetic enzymes presented as zinc-finger fusion proteins. Here we deliver the designs for an E. coli cell capable of synthesizing resveratrol using the Manifold platform. The Manifold platform will help lower costs and expand the applications of chemical biosynthesis.
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                 Our team is proud to share with iGEM our Resource Hub, a page dedicated to sharing all the best programs and services available for iGEM teams as they work to build their device and prepare for the competition. This idea began at the Mid-Atlantic Meetup when we chose to share with other teams some of the ways we were working online when not allowed in the lab. The response we got from this section of our presentation showed us that there was a lot of disconnect in the best ways to go about iGEM, so we decided to fill this need and create a hub for resources, based on function and accessibility. After talking to Nemanja Stijepovic and Leandros Tsiotos of the Video and Remote Technologies Committee about the perceived need for this resource, we created a plan to compile resources used and enjoyed by teams and compile the best into a single page. We presented our original list in the Global Meet Up, sharing the tools we had found most useful and how they allowed us to adapt to a summer spent outside the lab. Following this presentation, we distributed a survey where other iGEM teams could respond and tell us about the resources they found most useful. From the submissions and our own research, we built this document as a tool for all future iGEM teams to utilize. The survey is linked in the introduction, and we urge all teams to continue providing suggestions for expanding this list, and we hope future teams will find this hub useful in their endeavors.
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              The invention consists of a protein shell comprising one or more proteins, one or more nucleic acid scaffolds of which there can be multiple copies, anabolic and/or catabolic enzymes specific to the desired biosynthesis pathway each containing a nucleic acid binding domain, recognition sequences for the utilized nucleic acid binding domains, nucleic acid spacers, and a linkage between the nucleic acid scaffolds and the protein shell. The protein shell (10) can take the form of any closed or open surface that comprises one or more repeating protein units (12). Examples of valid shells include bacterial microcompartments such as the Pdu, Eut, and carboxysome microcompartments, as well as modified,  but not necessarily closed, surfaces composed of mutated versions of these microcompartment shell proteins. The nucleic acid scaffolds (18) comprise multiple recognition sequences (22) and spacers (32) and can be made from any form of nucleic acid, including: deoxyribonucleic acid, ribonucleic acid, and synthetic nucleic acids such as xeno nucleic acids and peptide nucleic acids among others. These scaffolds are attached to the protein shell. The pathway enzymes are biological proteins whose exact sequences are dependent on the given use case of the invention, but which all contain a nucleic acid binding domain either internal to their structure, or at their N or C terminus.
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              <b>Fig 1.</b> Figure taken from iGEM Tainan 2019 for demo purposes. Notice how the figure is much longer than it is wide, and two images are coupled together to achive this. Try to do that as well so it looks good.
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              Additionally, protein linkers are usually present between this nucleic acid binding domain and the enzyme structure to prevent inhibition of enzyme activity. However the exact linker(s) used, if any, is(are) also dependent on the specific use case of the invention. These pathway enzymes are attached to the nucleic acid scaffolds via their nucleic acid binding domains. The nucleic acid recognition sequences (22) are unique or semi-unique sequences of nucleic acid monomers on the nucleic acid scaffolds to which the utilized nucleic acid binding domains have some degree of molecular complementarity. These nucleic recognition sequences comprise most of the scaffold and mark the locations to which the DNA binding domains of the pathway enzymes attach to the scaffolds. The nucleic acid spacers (32) are relatively short sequences of nucleic acid monomers that are also present on the nucleic acid scaffolds, between the recognition sequences. The linkage between the nucleic acid scaffolds (18) and protein shell (10) provides a means by which the nucleic acid scaffolds are bound to the protein shell through direct or multi-molecule complementarity. This linkage is found between the nucleic acid scaffolds and the protein shell. One example is through the addition of a nucleic acid binding domain (24) to one or more of the shell proteins forming a nucleic acid binding domain, shell protein fusion (14). Like the pathway enzymes, this nucleic acid binding-domain can be either internal to the shell protein structure or at its N or C terminus, where the exact placement depends on the shell protein being utilized. Alternatively, one or more intermediate proteins can be used to adhere the nucleic acid scaffolds to the shell, where the region of the protein interacting with the shell binds the shell via protein-protein complementarity (28) with a given shell protein, and the region of the protein interacting with the nucleic acid scaffold binds another recognition sequence on the nucleic acid scaffold through another nucleic acid binding domain (30). This forms a shell protein binding, nucleic acid domain fusion (26).<br/><br/><br/>
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Revision as of 21:39, 24 October 2020

Manifold

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iGEM Resource Page
Our team is proud to share with iGEM our Resource Hub, a page dedicated to sharing all the best programs and services available for iGEM teams as they work to build their device and prepare for the competition. This idea began at the Mid-Atlantic Meetup when we chose to share with other teams some of the ways we were working online when not allowed in the lab. The response we got from this section of our presentation showed us that there was a lot of disconnect in the best ways to go about iGEM, so we decided to fill this need and create a hub for resources, based on function and accessibility. After talking to Nemanja Stijepovic and Leandros Tsiotos of the Video and Remote Technologies Committee about the perceived need for this resource, we created a plan to compile resources used and enjoyed by teams and compile the best into a single page. We presented our original list in the Global Meet Up, sharing the tools we had found most useful and how they allowed us to adapt to a summer spent outside the lab. Following this presentation, we distributed a survey where other iGEM teams could respond and tell us about the resources they found most useful. From the submissions and our own research, we built this document as a tool for all future iGEM teams to utilize. The survey is linked in the introduction, and we urge all teams to continue providing suggestions for expanding this list, and we hope future teams will find this hub useful in their endeavors.





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