The yeast enzyme Eht1 is an octanoyl-CoA:ethanol acyltransferase that also functions as a thioesterase.
Yeast
; 31(12): 463-74, 2014 Dec.
Article
em En
| MEDLINE
| ID: mdl-25308280
ABSTRACT
Fatty acid ethyl esters are secondary metabolites that are produced during microbial fermentation, in fruiting plants and in higher organisms during ethanol stress. In particular, volatile medium-chain fatty acid ethyl esters are important flavour compounds that impart desirable fruit aromas to fermented beverages, including beer and wine. The biochemical synthesis of medium-chain fatty acid ethyl esters is poorly understood but likely involves acyl-CoAethanol O-acyltransferases. Here, we characterize the enzyme ethanol hexanoyl transferase 1 (Eht1) from the brewer's yeast Saccharomyces cerevisiae. Full-length Eht1 was successfully overexpressed from a recombinant yeast plasmid and purified at the milligram scale after detergent solubilization of sedimenting membranes. Recombinant Eht1 was functional as an acyltransferase and, unexpectedly, was optimally active toward octanoyl-CoA, with k(cat) = 0.28 ± 0.02/s and K(M) = 1.9 ± 0.6 µm. Eht1 was also revealed to be active as a thioesterase but was not able to hydrolyse p-nitrophenyl acyl esters, in contrast to the findings of a previous study. Low-resolution structural data and site-directed mutagenesis provide experimental support for a predicted α/ß-hydrolase domain featuring a Ser-Asp-His catalytic triad. The S. cerevisiae gene YBR177C/EHT1 should thus be reannotated as coding for an octanoyl-CoAethanol acyltransferase that can also function as a thioesterase.
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1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Saccharomyces cerevisiae
/
Tioléster Hidrolases
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Acil Coenzima A
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Aciltransferases
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Proteínas de Saccharomyces cerevisiae
Tipo de estudo:
Prognostic_studies
Idioma:
En
Ano de publicação:
2014
Tipo de documento:
Article