Characterization of a regiospecific epivancosaminyl transferase GtfA and enzymatic reconstitution of the antibiotic chloroeremomycin
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Author
Lu, Wei
Oberthür, Markus
Leimkuhler, Catherine
Tao, Junhua
Kahne, Daniel
Walsh, Christopher T.
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https://doi.org/10.1073/pnas.0400277101Metadata
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Lu, W., M. Oberthur, C. Leimkuhler, J. Tao, D. Kahne, and C. T. Walsh. 2004. “Characterization of a Regiospecific Epivancosaminyl Transferase GtfA and Enzymatic Reconstitution of the Antibiotic Chloroeremomycin.” Proceedings of the National Academy of Sciences 101 (13): 4390–95. https://doi.org/10.1073/pnas.0400277101.Abstract
Chloroeremomycin, a vancomycin family glycopeptide antibiotic has three sugars, one D-glucose and two L-4-epi-vancosamines, attached to the crosslinked heptapeptide backbone by three glycosyltransferases, GtfA, -B, and -C. Prior efforts have revealed that GtfB and -C in tandem build an epivancosaminyl-1,2-glucosyl- disaccharide chain on residue 4 of the aglycone; however, the characterization of GtfA and glycosylation sequence of chloroeremomycin have been lacking. Here, we report the expression and purification of GtfA, as well as synthesis of its sugar donor, 2'-deoxy-thymidine 5'-diphosphate (dTDP)-beta-L-4-epi-vancosamine.. GtfA transfers 4-epi-vancosamine from the chemically synthesized dTDP-4-epi-vancosamine to the beta-OH-Tyr(6) residue of the aglycone, preferentially after GtfB action, to generate chloroorienticin B. With the preferred kinetic order of GtfB, then GtfA, then GtfC established, we have succeeded in reconstitution of chloroeremomycin from the heptapeptide aglycone by the sequential actions of the three enzymes.Terms of Use
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