Emopamil-binding protein, a mammalian protein that binds a series of structurally diverse neuroprotective agents, exhibits delta8-delta7 sterol isomerase activity in yeast.
Delta8-delta7 sterol isomerase is an essential enzyme on the sterol biosynthesis pathway in eukaryotes. This endoplasmic reticulum-resident membrane protein catalyzes the conversion of delta8-sterols to their corresponding delta7-isomers. No sequence data for high eukaryote sterol isomerase being available so far, we have cloned a murine sterol isomerase-encoding cDNA by functional complementation of the corresponding deficiency in the yeast Saccharomyces cerevisiae. The amino acid sequence deduced from the cDNA open reading frame is highly similar to human emopamil-binding protein (EBP), a protein of unknown function that constitutes a molecular target for neuroprotective drugs. A yeast strain in which the sterol isomerase coding sequence has been replaced by that of human EBP or its murine homologue recovers the ability to convert delta8-sterol into delta7-sterol, both in vivo and in vitro. In these recombinant strains, both cell proliferation and the sterol isomerization reaction are inhibited by the high affinity EBP ligand trifluoperazine, as is the case in mammalian cells but not in wild type yeast cell. In contrast, the recombinant strains are much less susceptible to the sterol inhibition effect of haloperidol and fenpropimorph, as compared with wild type yeast strains. Our results strongly suggest that EBP and delta8-delta7 sterol isomerase are identical proteins in mammals.
Silve
S
S
Department of Microbiology, Sanofi-Recherche, Labège Innopole BP137, F-31676 Labège Cédex, France.
Dupuy
P H
PH
Labit-Lebouteiller
C
C
Kaghad
M
M
Chalon
P
P
Rahier
A
A
Taton
M
M
Lupker
J
J
Shire
D
D
Loison
G
G
D000595:Amino Acid Sequence; D014150:Antipsychotic Agents / Q000378:metabolism / Q000494:pharmacology; D001483:Base Sequence; D015152:Blotting, Northern; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D002849:Chromatography, Gas; D002851:Chromatography, High Pressure Liquid; D018076:DNA, Complementary / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000235:genetics; D006220:Haloperidol / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D008969:Molecular Sequence Data; D009025:Morpholines / Q000378:metabolism / Q000494:pharmacology; D016366:Open Reading Frames; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D017386:Sequence Homology, Amino Acid; D013251:Steroid Isomerases / Q000378:metabolism*; D013261:Sterols / Q000096:biosynthesis; D015534:Trans-Activators / Q000235:genetics; D000071230:Transcriptional Regulator ERG; D014268:Trifluoperazine / Q000378:metabolism / Q000494:pharmacology
D014150:Antipsychotic Agents; D002352:Carrier Proteins; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; C072312:ERG protein, human; D009025:Morpholines; D013261:Sterols; D015534:Trans-Activators; D000071230:Transcriptional Regulator ERG; D014268:Trifluoperazine; C050435:fenpropimorph; D013251:Steroid Isomerases; C022669:delta(8)-delta(7)-sterol isomerase; C090445:EBP protein, human; D006220:Haloperidol