Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.27.3 (RNase T1)
 1,228 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cyclophilin and FK506 binding protein (FKBP) accelerate cis-trans peptidyl-prolyl isomerization and bind to and mediate the effects of the immunosuppressants cyclosporin A and FK506. The normal cellular functions of these proteins, however, are unknown. We altered the active sites of FKBP12 and mitochondrial cyclophilin from the yeast Saccharomyces cerevisiae by introducing mutations previously reported to inactivate these enzymes. Surprisingly, most of these mutant enzymes were biologically active in vivo. In accord with previous reports, all of the mutant enzymes had little or no detectable prolyl isomerase activity in the standard peptide substrate-chymotrypsin coupled in vitro assay. However, in a variation of this assay in which the protease is omitted, the mutant enzymes exhibited substantial levels of prolyl isomerase activity (5-20% of wild-type), revealing that these mutations confer sensitivity to protease digestion and that the classic in vitro assay for prolyl isomerase activity may be misleading. In addition, the mutant enzymes exhibited near wild-type activity with two protein substrates, dihydrofolate reductase and ribonuclease T1, whose folding is accelerated by prolyl isomerases. Thus, a number of cyclophilin and FKBP12 "active-site" mutants previously identified are largely active but protease sensitive, in accord with our findings that these mutants display wild-type functions in vivo. One mitochondrial cyclophilin mutant (R73A), and also the wild-type human FKBP12 enzyme, catalyze protein folding in vitro but lack biological activity in vivo in yeast. Our findings provide evidence that both prolyl isomerase activity and other structural features are linked to FKBP and cyclophilin in vivo functions and suggest caution in the use of these active-site mutations to study FKBP and cyclophilin functions.
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PMID:Functions of FKBP12 and mitochondrial cyclophilin active site residues in vitro and in vivo in Saccharomyces cerevisiae. 936 68

The in vitro protein folding activity of an FKBP (FK506 binding protein, abbreviated to MTFK) from a thermophilic archaeon, Methanococcus thermolithotrophicus, was investigated. MTFK exhibited FK506 sensitive PPIase (peptidyl prolyl cis-trans isomerase) activity which accelerated the speed of ribonuclease T1 refolding, which is rate-limited by isomerization of two prolyl peptide bonds. In addition, MTFK suppressed the aggregation of folding intermediates and elevated the final yield of rhodanese refolding. We called this activity of MTFK the chaperone activity. The chaperone activity of MTFK was also inhibited by FK506. Alignment of the amino acid sequences of MTFK with human FKBP12 showed that MTFK has two insertion sequences, consisting of 13 and 44 amino acids, at the N- and C-termini, respectively [Furutani, M., Iida, T., Yamano, S., Kamino, K., and Maruyama, T. (1998) J. Bacteriol. 180, 388-394]. To study the relationship between chaperone and PPIase activities of MTFK, mutant MTFKs with deletions of these insertion sequences or with amino acid substitutions were created. Their PPIase and chaperone activities were measured using a synthetic oligopeptide and denatured rhodanese as the substrates, respectively. The far-UV circular dichroism spectra of the wild type and the mutants were also analyzed. The results suggested that (1) the PPIase activity did not correlate with chaperone activity, (2) both insertion sequences were required for MTFK to take a proper conformation, and (3) the insertion sequence (44 amino acids) in the C-terminus was important for the chaperone activity.
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PMID:FK506 binding protein from a thermophilic archaeon, Methanococcus thermolithotrophicus, has chaperone-like activity in vitro. 1063 Oct 7