Activity-based metabolomic profiling of enzymatic function: Identification of Rv1248c as a mycobacterial 2-hydroxy-3-oxoadipate synthase Journal Article
| Authors: | de Carvalho, L. P. S.; Zhao, H.; Dickinson, C. E.; Arango, N. M.; Lima, C. D.; Fischer, S. M.; Ouerfelli, O.; Nathan, C.; Rhee, K. Y. |
| Article Title: | Activity-based metabolomic profiling of enzymatic function: Identification of Rv1248c as a mycobacterial 2-hydroxy-3-oxoadipate synthase |
| Abstract: | Activity based metabolomic profiling (ABMP) allows unbiased discovery of enzymatic activities encoded by genes of unknown function, and applies liquid-chromatography mass spectrometry (LC-MS) to analyze the impact of a recombinant enzyme on the homologous cellular extract as a physiologic library of potential substrates and products. The Mycobacterium tuberculosis protein Rv1248c was incompletely characterized as a thiamine diphosphate-dependent α-ketoglutarate decarboxylase. Here, recombinant Rv1248c catalyzed consumption of α-ketoglutarate in a mycobacterial small molecule extract with matched production of 5-hydroxylevulinate (HLA) in a reaction predicted to require glyoxylate. As confirmed using pure substrates by LC-MS, 1H-NMR, chemical trapping, and intracellular metabolite profiling, Rv1248c catalyzes C-C bond formation between the activated aldehyde of α-ketoglutarate and the carbonyl of glyoxylate to yield 2-hydroxy-3-oxoadipate (HOA), which decomposes to HLA. Thus, Rv1248c encodes an HOA synthase. © 2010 Elsevier Ltd. All rights reserved. |
| Keywords: | genetics; methodology; metabolism; enzymology; chembio; microbio; mycobacterium tuberculosis; nuclear magnetic resonance; lyase; nuclear magnetic resonance, biomolecular; 2 hydroxy 3 oxoadipate synthase; 2-hydroxy-3-oxoadipate synthase; metabolomics; oxo-acid-lyases |
| Journal Title: | Chemistry and Biology |
| Volume: | 17 |
| Issue: | 4 |
| ISSN: | 1074-5521 |
| Publisher: | Elsevier Inc. |
| Date Published: | 2010-04-23 |
| Start Page: | 323 |
| End Page: | 332 |
| Language: | English |
| DOI: | 10.1016/j.chembiol.2010.03.009 |
| PUBMED: | 20416504 |
| PROVIDER: | scopus |
| PMCID: | PMC2878197 |
| DOI/URL: | |
| Notes: | --- - "Cited By (since 1996): 6" - "Export Date: 20 April 2011" - "CODEN: CBOLE" - "Source: Scopus" |
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