title: The yeast cyclophilin multigene family: purification, cloning and characterization of a new isoform.
abstract: Cyclophilins (Cyps) constitute a highly conserved family of proteins present in a wide variety of organisms. Historically, Cyps were first identified by their ability to bind the immunosuppressive agent cyclosporin A (CsA) with high affinity; they later were found to have peptidyl-prolyl cis-trans isomerase (PPIase) activity, which catalyzes the folding of oligopeptides at proline-peptide bonds in vitro and may be important for protein folding in vivo. Cells of Saccharomyces cerevisiae contain at least two distinct Cyp-related PPIases encoded by the genes CYP1 and CYP2. A yeast strain (GL81) containing genomic disruptions of three known yeast PPIase-encoding genes [CYP1, CYP2 and RBP1 (for rapamycin-binding protein); Koltin et al., Mol. Cell. Biol. 11 (1991) 1718-1723] was previously constructed and found to be viable. Soluble fractions of these cells possess residual CsA-sensitive PPIase activity (2-5% of that present in wild-type cells as assayed in vitro). We have purified an approx. 18-kDa protein exhibiting PPIase activity from a soluble fraction of GL81 cells and determined that its N-terminal amino acid (aa) sequence exhibits significant homology (but nonidentity) to the Cyp1 and Cyp2 proteins. We designate the gene for this new protein, CYP3. Using a degenerate oligodeoxyribonucleotide (oligo) based on the N-terminal aa sequence, plus an internal oligo homologous to a conserved region within the portion of CYP1 and CYP2 that had been deleted in the genome, a CYP3-specific DNA fragment was generated by the polymerase chain reaction (PCR) using GL81 genomic DNA as a substrate. This PCR fragment was used as a probe to isolate CYP3 genomic and cDNA clones.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Gene Expression Sciences, SmithKline Beecham Pharmaceuticals, King of Prussia, PA 19406.
authors.forename: M M
authors.initials: MM
authors.lastname: McLaughlin
authors.forename: M J
authors.initials: MJ
authors.lastname: Bossard
authors.forename: P L
authors.initials: PL
authors.lastname: Koser
authors.forename: R
authors.initials: R
authors.lastname: Cafferkey
authors.forename: R A
authors.initials: RA
authors.lastname: Morris
authors.forename: L M
authors.initials: LM
authors.lastname: Miles
authors.forename: J
authors.initials: J
authors.lastname: Strickler
authors.forename: D J
authors.initials: DJ
authors.lastname: Bergsma
authors.forename: M A
authors.initials: MA
authors.lastname: Levy
authors.forename: G P
authors.initials: GP
authors.lastname: Livi
mesh_terms: D000591:Amino Acid Isomerases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D000595:Amino Acid Sequence; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D003001:Cloning, Molecular; D003524:Cyclosporins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D004271:DNA, Fungal; D008040:Genetic Linkage; D007527:Isoenzymes / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D005810:Multigene Family; D019696:Peptidylprolyl Isomerase; D016133:Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D016415:Sequence Alignment
chemical_list: D002352:Carrier Proteins; D003524:Cyclosporins; D004271:DNA, Fungal; D007527:Isoenzymes; D000591:Amino Acid Isomerases; D019696:Peptidylprolyl Isomerase