Ssp1, a site-specific parvulin homolog from Neurospora crassa active in protein folding.
Peptidyl-prolyl cis-trans-isomerases (PPIases) are enzymes capable of isomerizing a Xaa-Pro peptide bond. Three families of PPIases are known: cyclophilins, FKBPs, and parvulins. The physiological functions of the PPIases are only poorly understood. Eucaryotic members of the parvulin family have recently been shown to be essential for regulation of mitosis. Here we describe the purification and characterization of Ssp1, an abundant parvulin homolog from Neurospora crassa, which is unique among the known eucaryotic parvulins in containing a polyglutamine stretch between the N-terminal WW domain and the C-terminal PPIase domain. Ssp1 is a site-specific PPIase with respect to the amino acid N-terminal to the proline residue. Peptides with glutamate, phosphoserine, or phosphothreonine in the -1-position proved to be the best substrates. Ssp1 is not only able to isomerize small peptides but is also active in protein folding, as shown with mouse dihydrofolate reductase. Using the substrate specificity of Ssp1, we could identify Glu81-Pro82 as a PPIase-sensitive site in folding of dihydrofolate reductase. These results demonstrate that Ssp1 is a potent mediator of protein folding and that parvulins can serve as tools to elucidate rate-limiting steps in protein folding reactions.
Kops
O
O
Institut für Biochemie und Molekularbiologie der Universität Freiburg, Hermann-Herder-Strasse 7, D-79104 Freiburg, Germany.
Eckerskorn
C
C
Hottenrott
S
S
Fischer
G
G
Mi
H
H
Tropschug
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D015723:Gene Library; D006801:Humans; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D000072340:NIMA-Interacting Peptidylprolyl Isomerase; D009492:Neurospora crassa / Q000201:enzymology* / Q000235:genetics; D019696:Peptidylprolyl Isomerase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D017510:Protein Folding; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry / Q000235:genetics*
D017931:DNA Primers; D000072340:NIMA-Interacting Peptidylprolyl Isomerase; D011994:Recombinant Proteins; D013762:Tetrahydrofolate Dehydrogenase; C000606846:PIN1 protein, human; D019696:Peptidylprolyl Isomerase; C000606850:Pin1 protein, mouse