Structural control of chemoselectivity, stereoselectivity, and substrate specificity in membrane-bound fatty acid acetylenases and desaturases

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DOIResolve DOI: http://doi.org/10.1021/bi901605d
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TypeArticle
Journal titleBiochemistry
Volume48
Issue51
Pages1229812304; # of pages: 7
AbstractThe FAD2-like desaturases comprise a group of membrane-bound oxygenases involved in the modification of fatty acyl groups in plants and fungi. This group includes typical oleate desaturases which introduce a Δ12 cis double bond and more unusual enzymes such as Crep1, an acetylenase from the plant Crepis alpina, which introduces a triple bond in linoleate at the Δ12 position. In this study, the structure−function relationship between FAD2-like acetylenases and desaturases was examined through site-directed mutagenesis and heterologous expression. Eleven amino acid positions were identified that show complete evolutionary conservation within acetylenases or desaturases but have different amino acids in the other class of enzyme. Point mutants in Crep1 were constructed and expressed in yeast to test the role in fatty acid modification of the amino acids at the 11 positions. Results indicate the importance of five amino acid positions within Crep1 with regard to desaturase and acetylenase chemoselectivity, stereoselectivity, and substrate recognition. For example, relative to wild-type Crep1, the Y150F, F259L, and H266Q mutations all favored desaturation over acetylenation. The data indicate that small changes in primary sequence, particularly in the vicinity of the active site, can have profound changes on chemoselectivity and other aspects of the function of membrane-bound desaturase-like enzymes.
Publication date
LanguageEnglish
AffiliationNRC Plant Biotechnology Institute; National Research Council Canada
Peer reviewedYes
NRC number50155
NPARC number19073096
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Record identifier7b9e1d41-9aba-43a8-a7aa-5774d040bd2a
Record created2012-03-30
Record modified2016-05-09
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