A novel cytochrome b5-like domain is linked to the carboxyl terminus of the Saccharomyces cerevisiae delta-9 fatty acid desaturase.
Cytochrome b5 is an amphipathic mobile membrane protein that is predominantly located at the endoplasmic reticulum surface. It is an essential component of a number of membrane-bound redox systems. In animal and fungal cells cytochrome b5 is thought to be an electron donor for sterol modifying enzymes and fatty acid desaturases. Disruption of the Saccharomyces cytochrome b5 gene, however, yielded cells that had no nutritional requirement for either sterols or unsaturated fatty acids. Expression of sterol and fatty acid-modifying genes was increased in the cytochrome b5-disrupted cells, however, suggesting that cytochrome b5 may play some nonessential role in these functions. Unsaturated fatty acids in yeast are formed by Ole1p, an oxygen-dependent delta-9 fatty acid desaturase that is an intrinsic endoplasmic reticulum membrane protein. Although the yeast delta-9 fatty acid desaturase does not appear to require cytochrome b5, introduction of the rat liver stearoyl-CoA desaturase gene into an ole1-disrupted, cytochrome b5-disrupted yeast strain revealed that this enzyme specifically requires cytochrome b5 to function. Comparison of the coding sequences of the yeast and rat desaturase genes showed that the yeast protein contains a 113-amino acid carboxyl-terminal extension not found in the rat enzyme. That extension has regions of strong homology to cytochrome b5, particularly in the heme binding and electron transfer motifs. Truncation or disruption of the desaturase cytochrome b5-like domain in cells that contain the wild type diffusible b5 produced unsaturated fatty acid auxotrophy, suggesting that the cytochrome b5-like domain of Ole1p plays an essential role in the desaturase reaction.
Mitchell
A G
AG
Department of Biological Sciences, Rutgers University, Nelson Laboratories, Piscataway, New Jersey 08855-1059, USA.
Martin
C E
CE
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002417:Cattle; D016384:Consensus Sequence; D015786:Cytochromes b5 / Q000737:chemistry*; D017931:DNA Primers; D044943:Fatty Acid Desaturases / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics; D005838:Genotype; D008099:Liver / Q000201:enzymology; D008961:Models, Structural; D008969:Molecular Sequence Data; D010957:Plasmids; D016133:Polymerase Chain Reaction; D017433:Protein Structure, Secondary; D051381:Rats; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D013230:Stearoyl-CoA Desaturase / Q000096:biosynthesis / Q000737:chemistry
D017931:DNA Primers; D015786:Cytochromes b5; D044943:Fatty Acid Desaturases; D013230:Stearoyl-CoA Desaturase; C029627:delta-9 fatty acid desaturase