Structural and biochemical characterization of the cytosolic wheat cyclophilin TaCypA-1.
Cyclophilins belong to a family of proteins that bind to the immunosuppressive drug cyclosporin A (CsA). Several members of this protein family catalyze the cis-trans isomerization of peptide bonds preceding prolyl residues. The present study describes the biochemical and structural characteristics of a cytosolic cyclophilin (TaCypA-1) cloned from wheat (Triticum aestivum L.). Purified TaCypA-1 expressed in Escherichia coli showed peptidyl-prolyl cis-trans isomerase activity, which was inhibited by CsA with an inhibition constant of 78.3 nM. The specific activity and catalytic efficiency (kcat/Km) of the purified TaCypA-1 were 99.06 ± 0.13 nmol s(-1) mg(-1) and 2.32 × 10(5) M(-1) s(-1), respectively. The structures of apo TaCypA-1 and the TaCypA-1-CsA complex were determined at 1.25 and 1.20 Å resolution, respectively, using X-ray diffraction. Binding of CsA to the active site of TaCypA-1 did not result in any significant conformational change in the apo TaCypA-1 structure. This is consistent with the crystal structure of the human cyclophilin D-CsA complex reported at 0.96 Å resolution. The TaCypA-1 structure revealed the presence of a divergent loop of seven amino acids (48)KSGKPLH(54) which is a characteristic feature of plant cyclophilins. This study is the first to elucidate the structure of an enzymatically active plant cyclophilin which shows peptidyl-prolyl cis-trans isomerase activity and the presence of a divergent loop.