Catalytic mechanism and cofactor preference of dihydrodipicolinate reductase from methicillin-resistant Staphylococcus aureus
Details
Publication Year 2011-08-15, Volume 512, Issue #2, Page 167-174
Journal Title
ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS
Publication Type
Journal Article
Abstract
Given the rapid rise in antibiotic resistance, including methicillin resistance in Staphylococcus aureus (MRSA), there is an urgent need to characterize novel drug targets. Enzymes of the lysine biosynthesis pathway in bacteria are examples of such targets, including dihydrodipicolinate reductase (DHDPR, E.C. 1.3.1.26), which is the product of an essential bacterial gene. DHDPR catalyzes the NAD(P)H-dependent reduction of dihydrodipicolinate (DHDP) to tetrahydrodipicolinate (THDP) in the lysine biosynthesis pathway. We show that MRSA-DHDPR exhibits a unique nucleotide specificity utilizing NADPH (K-m = 12 mu M) as a cofactor more effectively than NADH (K-m = 26 mu M). However, the enzyme is inhibited by high concentrations of DHDP when using NADPH as a cofactor, but not with NADH. Isothermal titration calorimetry (ITC) studies reveal that MRSA-DHDPR has similar to 20-fold greater binding affinity for NADPH (K-d = 1.5 mu M) relative to NADH (K-d = 29 mu M). Kinetic investigations in tandem with ITC studies show that the enzyme follows a compulsory-order ternary complex mechanism; with inhibition by DHDP through the formation of a nonproductive ternary complex with NADP(+). This work describes, for the first time, the catalytic mechanism and cofactor preference of MRSA-DHDPR, and provides insight into rational approaches to inhibiting this valid antimicrobial target. (C) 2011 Elsevier Inc. All rights reserved.
Publisher
ELSEVIER SCIENCE INC
Keywords
X-RAY DIFFRACTION; ISOTHERMAL TITRATION CALORIMETRY; SINORHIZOBIUM-MELILOTI L5-30; ESCHERICHIA-COLI; MYCOBACTERIUM-TUBERCULOSIS; LYSINE BIOSYNTHESIS; 3-DIMENSIONAL STRUCTURE; BACILLUS-SUBTILIS; SYNTHASE; CRYSTALLIZATION
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Creation Date: 2011-08-15 12:00:00
Last Modified: 0001-01-01 12:00:00
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