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Biochemistry. 2007 Jan 16;46(2):359-68.

Characterization of the aminocarboxycyclopropane-forming enzyme CmaC.

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Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA.


The biosynthesis of the coronamic acid fragment of the pseudomonal phytotoxin coronatine involves construction of the cyclopropane ring from a gamma-chloro-L-allo-Ile intermediate while covalently tethered as a phosphopantetheinyl thioester to the carrier protein CmaD. The cyclopropane-forming catalyst is CmaC, catalyzing an intramolecular displacement of the gamma-Cl group by the alpha carbon. CmaC can be isolated as a Zn2+ protein with about 10-fold higher activity over the apo form. CmaC will not cyclize free gamma-chloro amino acids or their S-N-acetylcysteamine (NAC) thioester derivatives but will recognize some other carrier protein scaffolds. Turnover numbers of 5 min-1 are observed for Zn-CmaC, acting on gamma-chloro-L-aminobutyryl-S-CmaD, generating 1-aminocyclopropane-1-carbonyl (ACC)-S-CmaD. Products were detected either while still tethered to the phosphopantetheinyl prosthetic arm by mass spectrometry or after thioesterase-mediated release and derivatization of the free amino acid. In D2O, CmaC catalyzed exchange of one deuterium into the aminobutyryl moiety of the gamma-Cl-aminoacyl-S-CmaD, whereas the product ACC-S-CmaD lacked the deuterium, consistent with a competition for a gamma-Cl-aminobutyryl alpha-carbanion between reprotonation and cyclization. CmaC-mediated cyclization yielded solely ACC, resulting from C-C bond formation and no azetidine carboxylate from an alternate N-C cyclization. CmaC could cyclize gamma,gamma-dichloroaminobutyryl to the Cl-ACC product but did not cyclize delta- or epsilon-chloroaminoacyl-S-CmaD substrates.

[Indexed for MEDLINE]

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