1XPL

Crystal Structure of Staphylococcus aureus HMG-COA Synthase with Acetoacetyl-COA and Acetylated Cysteine

Summary for 1XPL

• Entry DOI: 10.2210/pdb1xpl/pdb
• Related: 1XPK 1XPM
• Descriptor: 3-hydroxy-3-methylglutaryl CoA synthase, SULFATE ION, ACETOACETYL-COENZYME A, ... (4 entities in total)
• Functional Keywords: hmg-coa synthase, hmgs, coenzyme a, thiolase fold, condensing enzyme, cholesterol biosynthesis, transferase
• Biological source: Staphylococcus aureus subsp. aureus
• Total number of polymer chains: 4
• Total formula weight: 181516.85

Authors

Theisen, M.J.,Misra, I.,Saadat, D.,Campobasso, N.,Miziorko, H.M.,Harrison, D.H.T. (deposition date: 2004-10-08, release date: 2004-11-02, Last modification date: 2024-11-20)

Primary citation

Theisen, M.J.,Misra, I.,Saadat, D.,Campobasso, N.,Miziorko, H.M.,Harrison, D.H.T.
3-hydroxy-3-methylglutaryl-CoA synthase intermediate complex observed in "real-time"
Proc.Natl.Acad.Sci.USA, 47:16442-16447, 2004

PubMed Abstract: The formation of carbon-carbon bonds via an acyl-enzyme intermediate plays a central role in fatty acid, polyketide, and isoprenoid biosynthesis. Uniquely among condensing enzymes, 3-hydroxy-3-methylglutaryl (HMG)-CoA synthase (HMGS) catalyzes the formation of a carbon-carbon bond by activating the methyl group of an acetylated cysteine. This reaction is essential in Gram-positive bacteria, and represents the first committed step in human cholesterol biosynthesis. Reaction kinetics, isotope exchange, and mass spectroscopy suggest surprisingly that HMGS is able to catalyze the "backwards" reaction in solution, where HMG-CoA is cleaved to form acetoacetyl-CoA (AcAc-CoA) and acetate. Here, we trap a complex of acetylated HMGS from Staphylococcus aureus and bound acetoacetyl-CoA by cryo-cooling enzyme crystals at three different times during the course of its back-reaction with its physiological product (HMG-CoA). This nonphysiological "backwards" reaction is used to understand the details of the physiological reaction with regards to individual residues involved in catalysis and substrate/product binding. The structures suggest that an active-site glutamic acid (Glu-79) acts as a general base both in the condensation between acetoacetyl-CoA and the acetylated enzyme, and the hydrolytic release of HMG-CoA from the enzyme. The ability to trap this enzyme-intermediate complex may suggest a role for protein dynamics and the interplay between protomers during the normal course of catalysis.

PubMed: 15498869
DOI: 10.1073/pnas.0405809101

Experimental method

X-RAY DIFFRACTION (2 Å)