A protein radical and ferryl intermediates are generated by linoleate diol synthase, a ferric hemeprotein with dioxygenase and hydroperoxide isomerase activities.
J Biol Chem
; 273(33): 20744-51, 1998 Aug 14.
Article
em En
| MEDLINE
| ID: mdl-9694817
ABSTRACT
Linoleate diol synthase (LDS) was isolated as a hemeprotein from the fungus Gaeumannomyces graminis. LDS converts linoleate sequentially to 8R-hydroperoxylinoleate (8-HPODE) through an 8-dioxygenase by insertion of molecular oxygen and to 7S,8S-dihydroxylinoleate through a hydroperoxide isomerase by intramolecular oxygen transfer. Light absorption and EPR spectra of LDS indicated that the heme iron was ferric and mainly high spin. Oxygen consumption during catalysis started after a short time lag which was reduced by 8-HPODE. Catalysis declined due to suicide inactivation. Stopped flow studies with LDS and 8-HPODE at 13 degreesC showed a rapid decrease in light absorption at 406 nm within 35 ms with a first order rate constant of 90-120 s-1. Light absorption at 406 nm then increased at a rate of approximately 4 s-1, whereas the absorption at 421 nm increased after a lag time of approximately 5 ms at a rate of approximately 70 s-1. EPR spectra at 77 K of LDS both with linoleic acid and 8-HPODE showed a transient doublet when quenched after incubation on ice for 3 s (major hyperfine splitting 2.3 millitesla; g = 2.005), indicating a protein radical. The relaxation properties of the protein radical suggested interaction with a metal center. 8-HPODE generated about twice as much radical as linoleic acid, and the 8-HPODE-induced radical appeared to be stable. Our results suggest that LDS may form, in analogy with prostaglandin H synthases, ferryl intermediates and a protein radical during catalysis.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Oxigenases
/
Prostaglandina-Endoperóxido Sintases
/
Oxirredutases Intramoleculares
/
Hemeproteínas
Idioma:
En
Revista:
J Biol Chem
Ano de publicação:
1998
Tipo de documento:
Article