INIBIOLP   05426
INSTITUTO DE INVESTIGACIONES BIOQUIMICAS DE LA PLATA "PROF. DR. RODOLFO R. BRENNER"
Unidad Ejecutora - UE
artículos
Título:
CYP52X1, representing a new cytochrome P450 subfamily, displays fatty acid hydroxylase activity and contributes to virulence and growth on insect cuticular substrates in the entomopathogenic fungus Beauveria bassiana
Autor/es:
SHIZHU ZHANG; EMILIE WIDEMANN; GRAUSEM BERNARD; AGNES LESOT; FRANCK PINOT; NICOLÁS PEDRINI; NEMAT O. KEYHANI
Revista:
JOURNAL OF BIOLOGICAL CHEMISTRY
Editorial:
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Referencias:
Lugar: Bethesda, Maryland; Año: 2012 vol. 287 p. 13477 - 13486
ISSN:
0021-9258
Resumen:
Infection
of insects by the entomopathogenic fungus Beauveria
bassiana proceeds via attachment and penetration of the host cuticle. The
outermost epicuticular layer or waxy layer of the insect represents a structure
rich in hydrocarbons including abundant amounts of alkanes and fatty acids. A
member of a novel cytochrome P450 subfamily, CYP52X1, implicated in hydrocarbon
assimilation by B. bassiana was
characterized. B. bassiana targeted gene knockouts lacking Bbcyp52x1 displayed reduced virulence when topically applied to Galleria mellonella, but no reduction in
virulence was noted when the insect cuticle was bypassed using an intrahemoceol
injection assay. No significant growth defects were noted in the mutant as
compared to the wild-type parent on any hydrocarbon substrates tested including
alkanes and fatty acids. Insect epicuticle germination assays, however, showed
reduced germination of DBbcyp52x1 conidia on grasshopper
wings as compared to the wild-type parent. Complementation of the gene-knock
with the full-length gene restored virulence and insect epicuticle germination
to wild-type levels. Heterologous expression of CYP52X1 in yeast was used to
characterize the substrate specificity of the enzyme. CYP52X1 displayed the
highest activity against midrange fatty acids (C12:0 and C14:0) and epoxy
stearic acid, 4-8 fold lower activity against C16:0, C18:1, and C18:2, and
little to no activity against C9:0 and C18:0. Mass spec analysis of the product
of the C16:0 reaction confirmed that the enzyme added a terminal hydroxyl to
the substrate (w-hydroxylase). These
data implicate CYP52X1 as contributing to the penetration of the host cuticle
via facilitating the assimilation of insect epicuticle hydrocarbons.