Reaction mechanism of 5,8-linoleate diol synthase, 10R-dioxygenase, and 8,11-hydroperoxide isomerase of Aspergillus clavatus.

Jernerén F, Garscha U, Hoffmann I, Hamberg M, Oliw EH

Biochim. Biophys. Acta 1801 (4) 503-507 [2010-04-00; online 2010-01-05]

Aspergilli express fusion proteins of an animal haem peroxidase domain with fatty acid dioxygenase (DOX) activity ( approximately 600 amino acids) and a functional or non-functional hydroperoxide isomerase/cytochrome P450 domain ( approximately 500 amino acids with EXXR and GPHXCLG motifs). 5,8-Linoleate diol synthases (LDS; ppoA) and 10R-DOX (ppoC) of Aspergillusnidulans and A. fumigatus belong to this group. Our objective was to determine the oxylipins formed from linoleic acid by A. clavatus and their mechanism of biosynthesis. A. clavatus oxidized linoleic acid to (8R)-hydroperoxylinoleic acid (8R-HPODE), (10R)-hydroperoxy-8(E),12(Z)-octadecadienoic acid (10R-HPODE), and to (5S,8R)-dihydroxy- and (8R,11S)-dihydroxylinoleic acids (DiHODE) as major products. This occurred by abstraction of the pro-S hydrogen at C-8 and antarafacial dioxygenation at C-8 or at C-10 with double bond migration. 8R-HPODE was then isomerized to 5S,8R-DiHODE and to 8R,11S-DiHODE by abstraction of the pro-S hydrogens at C-5 and C-11 of 8R-HPODE, respectively, followed by suprafacial oxygenation. The genome of A. clavatus codes for two enzymes, which can be aligned with >65% amino acid identity to 10R-DOX and 5,8-LDS, respectively. The 5,8-LDS homologue likely forms and isomerizes 8R-HPODE to 5S,8R-DiHODE. A third gene (ppoB) codes for a protein which carries a serine residue at the cysteine position of the P450 motif. This Cys to Ser replacement is known to abolish P450 2B4 catalysis and the hydroperoxide isomerase activity of 5,8-LDS, suggesting that ppoB of A. clavatus may not be involved in the biosynthesis of 8R,11S-DiHODE.

NGI Uppsala (Uppsala Genome Center)

National Genomics Infrastructure

PubMed 20045744

DOI 10.1016/j.bbalip.2009.12.012

Crossref 10.1016/j.bbalip.2009.12.012

pii: S1388-1981(09)00292-3

Publications 9.5.0