Production of Activated TDP-Deoxysugars in E. coli

DESCRIPTION provided by applicant The ultimate goal of the proposal presented herein is to use E coli as whole cell biocatalysts for the production of a wide variety of TDP deoxysugars including di and tri deoxysugars amino sugars and branched chain sugars These specialized activated hexoses are found as important structural components throughout plant and microbial secondary metabolites often playing a crucial role in conferring activity in bioactive natural products such as antibiotics and anticancer therapeutics In Phase I we proposed to investigate a novel approach to produce rare TDP deoxysugars in Escherichia coli through metabolic engineering During the Phase I study we successfully modified E coli AB to accumulate TDP keto deoxy D glucose TKDG a key intermediate of TDP deoxysugars through inactivations and overexpression of specific pathway enzymes Using exogenous TDP deoxysugar biosynthetic enzymes the accumulated TKDG pools were converted to specific TDP deoxysugars in yields up to mg L exceeding our Phase I goal The success of the feasibility study will be expanded in Phase II Specifically we will further increase TDP deoxysugar yields through additional genetic engineering experiments to limit loss of glucose phosphate to competing pathways increase thymidine triphosphates TTP pools reduce catabolite repression by glucose and optimize fermentation conditions We will also expand our range of TDP deoxysugar products by cloning and expressing several exogenous sugar biosynthetic genes for the production of additional complex TDP deoxysugars In Phase III we will commercialize the technology by carrying out Partnered Randamp D programs with strategic partners to make new glycoconjugates by offering TDP deoxysugars as well as deoxysugars as research reagents and by licensing the technology on an application specific basis PUBLIC HEALTH RELEVANCE This project is aimed toward developing E coli as a whole cell biocatalyst for the production of activated TDP deoxysugars These deoxysugars can be used to make derivatives of natural products with new therapeutic properties for example antibiotics that are effective against antibiotic resistant bacteria