R73A and H144Q mutants of the yeast mitochondrial cyclophilin Cpr3 exhibita low prolyl isomerase activity in both peptide and protein-folding assays

Previously we reported that the R73A and H144Q variants of the yeast cyclop hilin Cpr3 mere virtually inactive in a protease-coupled peptide assay, but retained activity as catalysts of a proline-limited protein folding reacti on [Scholz, C, et al, (1997) FEES Lett, 414, 69-73], A reinvestigation reve aled that in fact these two mutations strongly decrease the prolyl isomeras e activity of Cpr3 in both the peptide and the protein-folding assay. The h igh folding activities found previously originated from a contamination of the recombinant Cpr3 proteins with the Escherichia coli protein SlyD, a pro lyl isomerase that co-purifies with His-tagged proteins. SlyD is inactive i n the peptide assay, but highly active in the protein-folding assay. (C) 19 99 Federation of European Biochemical Societies.