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                3) Four path-scoring criteria
 
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The main objective of our project is to construct a novel “protein whip” platform, with which we can make <i>Corynebacterium glutamicum</i> to express other corynebacterium’s pili structure comprised of chains of a protein of our choice. As our first try, we decided to make pili made out of green fluorescence proteins (GFP); in order to do so, we substituted SpaA protein, one of the surface proteins in the Pilin A gene cluster, into green fluorescence protein, and transformed a vector containing the modified Pilin A gene cluster into a <i>C. glutamicum</i> strain. <br />Our “protein whip” platform is expected to have many practical applications. For example, pili made out of an enzyme, enzyme whip will enable the reaction to take place with high efficiency, for a great number of the enzyme included in the pili will be able to “attack” the reactants simultaneously. Biofilms made of strains of bacteria that express pili comprised of chains of specific amino acids such as histidine or cysteine that readily bind to heavy metals may be utilized to purify water contaminated with heavy metals. <br />
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① NADH and NADPH
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Having a number of potential applications is not the sole merit of our project; by using <i>C. glutamicum</i> instead of widely exploited <i>Escherichia coli</i>, our project also contributes to expanding model organisms used in synthetic biology beyond <i>E. coli</i>.
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Like an ATP (adenosine triphosphate) which serves as a convenient and versatile storage in cells, NADH (nicotinamide adenine dinucleotide) and NADPH (nicotinamide adenine dinucleotide phosphate) serve as activated carriers specialized in carrying high-energy electrons and hydrogen atoms (see Figure 1-a).  
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Revision as of 15:18, 18 September 2015

1) KEGG database

The program ‘Gil’ is a project based on KEGG (Kyoto Encyclopedia of Genesand Genomes) database (Kanehisa M., et al. 2014; and Kanehisa M., Goto S., 2000). Its information were accumulated by genome sequencing and other biological research. It offers well-combined metabolic-related pathways. Also we organized and parsed these database that provide compound, reaction, gene and enzymatic information. Especially, we built RPair, a main network in four reaction pair groups. However, there are some shortcomings in KEGG database. First, it doesn't provide thermodynamic information that is a crucial criterion to determine reaction direction. Though we cannot obtain thermodynamic information in KEGG database, we tried to compensate it by utilizing Equilibrater and anotherdatabase. Additionally KEGG database, however, show reactions in bidirectional way in which usually do not process as it is a thermodynamically unfavorable reaction. Lastly, every glycan in KEGG database is not connected with compound. By origin, glycans have G number (ex. G00000) but there are some cases that glycans have C number (ex. C00000) that kegg compounds have. Specially, they can react to compounds or glycans. However, many glycans don’t have C number, so they can not react with compounds and they even don’t have any reactions in glycans. For example, glycan named neoagarobiose in the path that has been being studying for experimental validation doesn’t have C number. So we added the information of the glycan found other databases such as “-------”* and -------** to data obtained from kegg compounds to complement kegg’s fault. * a database of chemical molecules maintained by the National Center for Biotechnology information (NCBI) (Wikipedia) ** Eun Ju Yun et al. (2015), The novel catabolic pathway of 3,6-anhydro-Lgalactose, the main component of red macroalgae, in a marine bacterium, Environmental Microbiology, 1677 – 1688p We used K12 as our E. coli model strain. It is the most commonly used E. coli strain in biological lab.

2) Thermodynamic information

Thermodynamic constrain is one of the most important factors of biological experiments. Therefore, the program ‘Gil’ provides Gibbs energy of each metabolic reaction. The program ‘Gil’ obtained thermodynamic datafrom two separated databases even though they were obtained from the same program, eQuilibrator (Flamholz A., et al. 2012). The first database is composed of Gibbs energy from standard condition—pH 7.0 and 0.1 M ionic strength. These data are correct, but the size of the database is smaller than the other one due to the lack of wetlab experiments. On the other hand, the second version is made up of values from calculation. This database has much more values than the first one, however it is hard for researchers to apply this values directly to their experiments since they are not experimentally proved. Thus, additional attention is needed to utilize the second version of data.

Thermodynamic constrain is one of the most important factors of biological experiments. Therefore, the program ‘Gil’ provides Gibbs energy of each metabolic reaction. The program ‘Gil’ obtained thermodynamic datafrom two separated databases even though they were obtained from the same program, eQuilibrator (Flamholz A., et al. 2012). The first database is composed of Gibbs energy from standard condition—pH 7.0 and 0.1 M ionic strength. These data are correct, but the size of the database is smaller than the other one due to the lack of wetlab experiments. On the other hand, the second version is made up of values from calculation. This database has much more values than the first one, however it is hard for researchers to apply this values directly to their experiments since they are not experimentally proved. Thus, additional attention is needed to utilize the second version of data.
3) Four path-scoring criteria

① NADH and NADPH Like an ATP (adenosine triphosphate) which serves as a convenient and versatile storage in cells, NADH (nicotinamide adenine dinucleotide) and NADPH (nicotinamide adenine dinucleotide phosphate) serve as activated carriers specialized in carrying high-energy electrons and hydrogen atoms (see Figure 1-a).