Histidine 109 in peptidyl-prolyl cis-trans isomerase of Bacillus subtilis plays an important role in catalysis and in cyclosporin A binding

The cyclophilin of Bacillus subtilis has a moderate affinity to cyclosporin A (IC 50: 120 nM) and low catalytic activity ( k cat/ K m: 1.1 μM −1 s −1) when compared to other ubiquitous peptidyl-prolyl cis-trans isomerases (PPIases). The active site residues V52, H90 and H109, which are not conserved...

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Bibliographic Details
Published in:FEMS microbiology letters 1997-09, Vol.154 (1), p.139-144
Main Authors: Achenbach, Tatjana V., Göthel, Sven F., Marahiel, Mohamed A.
Format: Article
Language:English
Subjects:
Bacillus subtilis
Bacillus subtilis - chemistry
Bacillus subtilis - enzymology
Bacillus subtilis - genetics
Bacterial Proteins - chemistry
Bacterial Proteins - genetics
Bacterial Proteins - metabolism
Cyclophilin
Cyclosporin A
Cyclosporine - metabolism
Gene Expression Regulation, Bacterial
Histidine - analysis
Hydrogen-Ion Concentration
Molecular Sequence Data
Mutagenesis
Peptidyl-prolyl cis-trans isomerase
Peptidylprolyl Isomerase - chemistry
Peptidylprolyl Isomerase - genetics
Peptidylprolyl Isomerase - metabolism
Protein Binding - physiology
Sequence Homology, Amino Acid
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Summary:The cyclophilin of Bacillus subtilis has a moderate affinity to cyclosporin A (IC 50: 120 nM) and low catalytic activity ( k cat/ K m: 1.1 μM −1 s −1) when compared to other ubiquitous peptidyl-prolyl cis-trans isomerases (PPIases). The active site residues V52, H90 and H109, which are not conserved within other peptidyl-prolyl cis-trans isomerases, were found to play an important role in cyclosporin A binding and catalytic activity. In this work we report on double mutations of these residues, which greatly improved cyclosporin A affinity and catalytic activity. The H90N/H109W mutation displayed an IC 50 value of 46 nM whereas the V52M/H109F mutation exhibited over 18-fold higher catalytic activity than that detected for wild-type PPIase. The mutations H109W and H109F of the B. subtilis PPIase showed no change in cyclosporin A affinity and catalytic activity between pH 6 and 8. In contrast, wild-type PPIase (H109) showed up to 10-fold reduction below pH 7.5, both in cyclosporin A affinity and in catalytic activity. These findings clearly underline the importance of the unique H109 residue in the B. subtilis enzyme.
ISSN:0378-1097
1574-6968
DOI:10.1016/S0378-1097(97)00314-5