Author
Awofiranye, Adeola, Esther
Other Contributors
Koffas, Mattheos, AG; Bystroff, Chris; Wang, Chunyu; Colon, Wilfredo;
Date Issued
2022-12
Subject
Biology
Degree
PhD;
Terms of Use
This electronic version is a licensed copy owned by Rensselaer Polytechnic Institute (RPI), Troy, NY. Copyright of original work retained by author.;
Abstract
We metabolically engineered E. coli strains to produce chondroitin sulfate (CS). We then used these engineered strains to produce derivatives of CS through feeding experiments. We Synthesized N-glycolyl chondroitin (Gc-CN) and N-glycolyl chondroitin sulfate (Gc-CS). These derivatives have medical and evolutionary applications. Gc-CN/Gc-CS is hypothesized to be able to detect diseases like carcinomas, atherosclerosis, etc. because it is a metabolite of the sialic acids that contributes to these diseases in humans, N-glycolylneuraminic acid (Neu5Gc). Furthermore, the derivatives can help date the loss of the CMAH gene; a gene lost after a last common ancestor with the great apes. We successfully synthesized Gc-CN and Gc-CS in metabolically engineered E. coli K4 adapted for CS production. We fed the bacteria with a glucose carbon source supplemented with chemically synthesized N-glycolyl glucosamine (GlcNGc), which allowed the incorporation of the N-glycolyl into chondroitin. After exploring the pathway for chondroitin sulfate synthesis, we chose to increase N-glycolyl incorporation into chondroitin. To do this, we investigated the effect of downregulating some genes in order to shift the metabolic flux towards N-glycolyl incorporation. We investigated the effect of knocking the bifunctional N-acetylglucosamine-1-phosphate uridyltransferase and glucosamine-1-phosphate acetyltransferase gene (GlmU) and the Glutamine-fructose-6-phosphate aminotransferase gene (GlmS). We Further decided to synthesize other chondroitin derivatives by isotopically labelling them. Using the same metabolically engineered E. coli strain for CS production, we produced deuterated chondroitin and 13C labeled.;
Description
December2022; School of Science
Department
Dept. of Biological Sciences;
Publisher
Rensselaer Polytechnic Institute, Troy, NY
Relationships
Rensselaer Theses and Dissertations Online Collection;
Access
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Attribution-Noncommercial-No Derivative Works 3.0 license. No commercial use or derivatives
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