Campus Units

Biochemistry, Biophysics and Molecular Biology

Document Type

Article

Publication Version

Published Version

Publication Date

2008

Journal or Book Title

Journal of Bacteriology

Volume

190

Issue

14

First Page

4888

Last Page

4893

DOI

10.1128/JB.00454-08

Abstract

Biotin-containing 3-methylcrotonyl coenzyme A (MC-CoA) carboxylase (MCCase) and geranyl-CoA (G-CoA) carboxylase (GCCase) from Pseudomonas aeruginosa were expressed as His-tagged recombinant proteins in Escherichia coli. Both native and recombinant MCCase and GCCase showed pH and temperature optima of 8.5 and 37°C. The apparent K0.5 (affinity constant for non-Michaelis-Menten kinetics behavior) values of MCCase for MC-CoA, ATP, and bicarbonate were 9.8 μM, 13 μM, and 0.8 μM, respectively. MCCase activity showed sigmoidal kinetics for all the substrates and did not carboxylate G-CoA. In contrast, GCCase catalyzed the carboxylation of both G-CoA and MC-CoA. GCCase also showed sigmoidal kinetic behavior for G-CoA and bicarbonate but showed Michaelis-Menten kinetics for MC-CoA and the cosubstrate ATP. The apparent K0.5 values of GCCase were 8.8 μM and 1.2 μM for G-CoA and bicarbonate, respectively, and the apparent Km values of GCCase were 10 μM for ATP and 14 μM for MC-CoA. The catalytic efficiencies of GCCase for G-CoA and MC-CoA were 56 and 22, respectively, indicating that G-CoA is preferred over MC-CoA as a substrate. The enzymatic properties of GCCase suggest that it may substitute for MCCase in leucine catabolism and that both the MCCase and GCCase enzymes play important roles in the leucine and acyclic terpene catabolic pathways.

Comments

This is an article from Journal of Bacteriology 190 (2008): 4888, doi:10.1128/JB.00454-08. Posted with permission.

Copyright Owner

American Society for Microbiology

Language

en

File Format

application/pdf

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