Abstract
The JC201 strain of Eschericia coli contains a temperature-sensitive lesion in lysophosphatidate acyltransferase (LPAT) activity. The LPAT gene from JC201 was isolated by PCR and a single mutant nucleotide, adenine-440, was identified by DNA sequence analysis. Site-directed mutagenesis converted the mutant adenine-440 back to the native guanine-440 nucleotide. The restored LPAT gene rescued JC201 cells at the non-permissive temperature. The fatty acid substrate specificity of LPAT from Eschericia coli was altered by site-directed mutagenesis of a single amino acid in the restored LPAT gene. Threonine-122 of LPAT was changed to alanine or leucine. A change from threonine-122 to alanine increased the substrate specificity in vitro for oleoyl-CoA and linoleoyl-CoA; whereas a change to leucine increased the substrate specificity for lignoceroyl-CoA.
Original language | English (US) |
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Pages (from-to) | 79-84 |
Number of pages | 6 |
Journal | Biochemical and Biophysical Research Communications |
Volume | 244 |
Issue number | 1 |
DOIs | |
State | Published - Mar 6 1998 |
ASJC Scopus subject areas
- Biochemistry
- Biophysics
- Molecular Biology