Enzymatic catalysis of prolyl isomerization in an unfolding protein.
Prolyl isomerases are able to accelerate slow steps in protein refolding that are limited in rate by cis/trans isomerizations of Xaa-Pro peptide bonds. We show here that prolyl isomerizations in the course of protein unfolding are also well catalyzed. To demonstrate catalysis we use cytoplasmic prolyl isomerase from Escherichia coli as the enzyme and reduced and carboxymethylated ribonuclease T1 as the substrate. This form of ribonuclease T1 without disulfide bonds is nativelike folded only in the presence of moderate concentrations of NaCl. Unfolding can be induced by reducing the NaCl concentration at ambient temperature and in the absence of denaturants. Under these conditions prolyl isomerase retains its activity and it catalyzes prolyl cis/trans isomerization in the unfolding protein. Under identical conditions within the NaCl-induced transition unfolding and refolding are catalyzed with equal efficiency. The stability of the protein and thus the final distribution of unfolded and folded molecules attained at equilibrium is unchanged in the presence of prolyl isomerase. These results demonstrate that prolyl isomerase functions in protein folding as an enzyme and catalyzes prolyl isomerization in either direction.
Mücke
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Laboratorium für Biochemie, Universität Bayreuth, Germany.
Schmid
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D000591:Amino Acid Isomerases / Q000378:metabolism*; D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000378:metabolism*; D002384:Catalysis; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology; D007700:Kinetics; D008745:Methylation; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D019696:Peptidylprolyl Isomerase; D011487:Protein Conformation; D006163:Ribonuclease T1 / Q000737:chemistry / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology; D013053:Spectrophotometry
D002352:Carrier Proteins; D012965:Sodium Chloride; D006163:Ribonuclease T1; D000591:Amino Acid Isomerases; D019696:Peptidylprolyl Isomerase