Isolation and Amino Acid Sequence of a New 22-kDa FKBP-like Peptidyl-prolyl cis/trans-Isomerase of Escherichia coli

Abstract

We identified a periplasmic peptidyl-prolyl cis/trans-isomerase (PPIase) of the (FK506-binding protein (FKBP) type in Escherichia coli (FK506 represents a natural peptidomacrolide containing an acylated pipe-colic acid residue). After purification to homogeneity, its complete amino acid sequence was determined by a combination of Edman degradation and electrospray mass spectrometry of the authentic protein and pep-tides generated by proteolysis. The molecular mass calculated from the amino acid sequence of the protein was 22,085.53 Da, which corresponded perfectly with the value of 22,084 1.47 Da as determined by mass spec-trometry. The corresponding gene was cloned and analyzed , and Southern blot experiments revealed the existence of similar genes in various Gram-negative bacteria. The amino acid sequence of the novel FKBP22 shows similarity to Mip (macrophage infectivity poten-tiator)-like proteins produced by a number of patho-genic bacteria. However, FKBP22 is inhibited more strongly by FK506 than are other Mip-homologues, as indicated by the K i value of 25 nM. The subsite specificity regarding the P 1 position of the substrate resembles that for Mip-FKBP25 from Legionella pneumophila. The mature FKBP22 enzyme of 205 amino acids exists as a dimer in solution.