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Title: Directed mutagenesis alters the stereochemistry of catalysis by isolated ketoreductase domains from the erythromycin polyketide synthase.
Authors: Baerga-Ortiz, A
Popovic, B
Siskos, AP
O'Hare, HM
Spiteller, D
Williams, MG
Campillo, N
Spencer, JB
Leadlay, PF
First Published: Mar-2006
Citation: CHEM BIOL, 2006, 13 (3), pp. 277-285
Abstract: The ketoreductase (KR) domains eryKR(1) and eryKR(2) from the erythromycin-producing polyketide synthase (PKS) reduce 3-ketoacyl-thioester intermediates with opposite stereospecificity. Modeling of eryKR(1) and eryKR(2) showed that conserved amino acids previously correlated with production of alternative alcohol configurations lie in the active site. eryKR(1) domains mutated at these positions showed an altered stereochemical outcome in reduction of (2R, S)-2-methyl-3-oxopentanoic acid N-acetylcysteamine thioester. The wild-type eryKR(1) domain exclusively gave the (2S, 3R)-3-hydroxy-2-methylpentanoic acid N-acetylcysteamine thioester, while the double mutant (F141W, P144G) gave only the (2S, 3S) isomer, a switch of the alcohol stereochemistry. Mutation of the eryKR(2) domain, in contrast, greatly increased the proportion of the wild-type (2R, 3S)-alcohol product. These data confirm the role of key residues in stereocontrol and suggest an additional way to make rational alterations in polyketide antibiotic structure.
DOI Link: 10.1016/j.chembiol.2006.01.004
ISSN: 1074-5521
Type: Journal Article
Appears in Collections:Published Articles, Dept. of Infection, Immunity and Inflammation

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