Difference between revisions of "Team:Korea U Seoul/Project/Biological Background/content"
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<img src="https://static.igem.org/mediawiki/2015/4/43/KoreaUSeoul_BiologicalBackground_Figure1.jpg"> | <img src="https://static.igem.org/mediawiki/2015/4/43/KoreaUSeoul_BiologicalBackground_Figure1.jpg"> | ||
| − | < | + | <p><Figure 1></p> |
<p> The NAD + and NADH molecules are indistinguishable in structure to NADP + and NADPH, respectively, except the fact that the phosphate group is absent from the former. However, this smalldifference is crucial for their distinctive roles. Although the extra phosphate group on NADPH is located far away from the region involved in electron transfer (see Figure 1-b), it gives the molecule a slight difference in its shape from that of NADH, so NADPH and NADH bind to different sets of enzymes as substrates. Thus, thetwo types of carriers are used to transfer electrons (or hydride ions) between different sets of molecules. This role separationis crucial for organisms to regulate two sets of electron-transfer reactions independently. A NADPH operates mainly with enzymes that catalyze anabolic reactions, providing the high-energy electrons needed to synthesize energy-rich biological molecules. In contrast, a NADH has a special role as an intermediate in the catabolic system of reactions that generate ATP through the oxidation of compounds (Alberts, B., et al. 2002). | <p> The NAD + and NADH molecules are indistinguishable in structure to NADP + and NADPH, respectively, except the fact that the phosphate group is absent from the former. However, this smalldifference is crucial for their distinctive roles. Although the extra phosphate group on NADPH is located far away from the region involved in electron transfer (see Figure 1-b), it gives the molecule a slight difference in its shape from that of NADH, so NADPH and NADH bind to different sets of enzymes as substrates. Thus, thetwo types of carriers are used to transfer electrons (or hydride ions) between different sets of molecules. This role separationis crucial for organisms to regulate two sets of electron-transfer reactions independently. A NADPH operates mainly with enzymes that catalyze anabolic reactions, providing the high-energy electrons needed to synthesize energy-rich biological molecules. In contrast, a NADH has a special role as an intermediate in the catabolic system of reactions that generate ATP through the oxidation of compounds (Alberts, B., et al. 2002). | ||
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<p>First, in the case of NADPH, it is the key for determining the efficiency of certain biosynthetic pathways. The oxidation during a pentose phosphorylation produce most NADPH which is needed in the biosynthesis of certain metabolites. Therefore, thecarbon flow of the pentose phosphorylation path is considered to be the targets of metabolic engineering. For example, microorganisms found in the Corynebacterium genus, having enhanced L-ornithine production, not only activate pentose phosphate pathway but also increase the specific activities of G6PDH and 6PGD, which are enzymes related to the pentose phosphate pathway, by reducing or inactivating intrinsic GDH (glucose dehydrogenase) (see Figure 2). Consequently, the improved NADPH production makes L-ornithine yield to increase, which may have the possibility of being widely utilized for efficient and economical applications (Cho J., Ko Y. 2012). To summarize, if we find a path with improved NADPH yield, we could get more products in other relevant paths. | <p>First, in the case of NADPH, it is the key for determining the efficiency of certain biosynthetic pathways. The oxidation during a pentose phosphorylation produce most NADPH which is needed in the biosynthesis of certain metabolites. Therefore, thecarbon flow of the pentose phosphorylation path is considered to be the targets of metabolic engineering. For example, microorganisms found in the Corynebacterium genus, having enhanced L-ornithine production, not only activate pentose phosphate pathway but also increase the specific activities of G6PDH and 6PGD, which are enzymes related to the pentose phosphate pathway, by reducing or inactivating intrinsic GDH (glucose dehydrogenase) (see Figure 2). Consequently, the improved NADPH production makes L-ornithine yield to increase, which may have the possibility of being widely utilized for efficient and economical applications (Cho J., Ko Y. 2012). To summarize, if we find a path with improved NADPH yield, we could get more products in other relevant paths. | ||
</p> | </p> | ||
| − | <img src="https://static.igem.org/mediawiki/2015/f/f6/KoreaUSeoul_BiologicalBackground_Figure2.png" width="700" height=" | + | <img src="https://static.igem.org/mediawiki/2015/f/f6/KoreaUSeoul_BiologicalBackground_Figure2.png" width="700" height="800"> |
| − | < | + | <p><Figure 2></p> |
| − | <p> | + | <p>In the same context, if theNADH yield is increased, the efficiency of the path using the NADH will also be enhanced. In addition, cells grown aerobically may possibly use the extra NADH for ATP generation through the electron transport system. According to the research on the availability of intracellular NADH (Berríos-Rivera SJ., Bennett GN, San KY 2002), the higher NADH availability increased dramatically the ethanol to acetate (Et/Ac) ratio by 27-fold. The cause of this amazing increase is proven by an experiment comparing BS1 (pSBF2), which is inactivated by the native FDH (formate dehydrogenase) that regenerates NADH, with GJT001 (pDHK29), the control, under anaerobic conditions. |
</p> | </p> | ||
<p>For these reasons, our program suggests the paths in the order of the amount of NADPH and NADH production. Users will be able to get efficient and productive paths with the information provided by our program, with less effort. | <p>For these reasons, our program suggests the paths in the order of the amount of NADPH and NADH production. Users will be able to get efficient and productive paths with the information provided by our program, with less effort. | ||
Revision as of 16:28, 18 September 2015
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.
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.
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).
The NAD + and NADH molecules are indistinguishable in structure to NADP + and NADPH, respectively, except the fact that the phosphate group is absent from the former. However, this smalldifference is crucial for their distinctive roles. Although the extra phosphate group on NADPH is located far away from the region involved in electron transfer (see Figure 1-b), it gives the molecule a slight difference in its shape from that of NADH, so NADPH and NADH bind to different sets of enzymes as substrates. Thus, thetwo types of carriers are used to transfer electrons (or hydride ions) between different sets of molecules. This role separationis crucial for organisms to regulate two sets of electron-transfer reactions independently. A NADPH operates mainly with enzymes that catalyze anabolic reactions, providing the high-energy electrons needed to synthesize energy-rich biological molecules. In contrast, a NADH has a special role as an intermediate in the catabolic system of reactions that generate ATP through the oxidation of compounds (Alberts, B., et al. 2002).
Then, why should we focus on the larger amount of NADH and NADPH as products?
First, in the case of NADPH, it is the key for determining the efficiency of certain biosynthetic pathways. The oxidation during a pentose phosphorylation produce most NADPH which is needed in the biosynthesis of certain metabolites. Therefore, thecarbon flow of the pentose phosphorylation path is considered to be the targets of metabolic engineering. For example, microorganisms found in the Corynebacterium genus, having enhanced L-ornithine production, not only activate pentose phosphate pathway but also increase the specific activities of G6PDH and 6PGD, which are enzymes related to the pentose phosphate pathway, by reducing or inactivating intrinsic GDH (glucose dehydrogenase) (see Figure 2). Consequently, the improved NADPH production makes L-ornithine yield to increase, which may have the possibility of being widely utilized for efficient and economical applications (Cho J., Ko Y. 2012). To summarize, if we find a path with improved NADPH yield, we could get more products in other relevant paths.
In the same context, if theNADH yield is increased, the efficiency of the path using the NADH will also be enhanced. In addition, cells grown aerobically may possibly use the extra NADH for ATP generation through the electron transport system. According to the research on the availability of intracellular NADH (Berríos-Rivera SJ., Bennett GN, San KY 2002), the higher NADH availability increased dramatically the ethanol to acetate (Et/Ac) ratio by 27-fold. The cause of this amazing increase is proven by an experiment comparing BS1 (pSBF2), which is inactivated by the native FDH (formate dehydrogenase) that regenerates NADH, with GJT001 (pDHK29), the control, under anaerobic conditions.
For these reasons, our program suggests the paths in the order of the amount of NADPH and NADH production. Users will be able to get efficient and productive paths with the information provided by our program, with less effort.