Genetic Manipulation of the Fatty Acid Chain Length Pattern in Yeast

✍ Scribed by Hühne, Kathrin ;Schweizer, M. ;Schweizer, E.

• Publisher: John Wiley and Sons
• Year: 1990
• Weight: 620 KB
• Volume: 92
• Category: Article
• ISSN: 0931-5985
• DOI: 10.1002/lipi.19900920606

✦ Synopsis

Full length fatty acyl thioesterase I1 cDNA was isolated from a rat mammary gland cDNA library. The coding sequence of the enzyme was placed under the control of the yeast ADHl promoter by insertion into the expression vector pVl-U. From this construction, pRThII, the mt thioesterase I1 gene was expressed in yeast at both the transcriptional and translational levels. The plasmid-encoded enzyme synthesized in pRThIItransformed yeast cells was purified about 190 fold up to a fmal specific activity of 834 units per milligram. Upon SDS gel electrophoresis, the purified enzyme was resolved into two main components with molecular weights of 28000 and 29000 daltons, respectively, both of which crossreacted with a specific anti-thioesterase I1 antiserum. Only traces of a third, immunologically unrelated protein band could be detected in the gel. The size of the larger thioesterase band corresponds well with the molecular weight (29.4 kDa) theoretically expected from the nucleotide sequence of the gene.