YEAST CYTOSOLIC CYCLOPHILIN A AS ANALOGY OF HUMAN ISOFORM. Michiko Konno, Sawako Fujioka, Akiko Kashima, Ochanomizu University, Department of Chemistry, Faculty of Science, Otsuka, bunkyo-ku, Tokyo 112 Japan
Prokaryotes to eukaryotes have ubiquitously and abundantly cyclophilin (CyP) soluble in the cytoplasm; CyPA and membrane-bound CyP; CyPB. While CyPB is seen to be homolog to ninaA, biological role of CyPA is still poorly understood. Even though yeast strains lacking either or both CyPs have been reported to grow normally, CyPA should be involved in some important machinery for life cycle of alive cell. Close comparison of structrural natures between yeast CyPA here determined and human CyPA (hCyPA) previously reported (1) is made to confirm functional identity of these two.
Crystals belong to triclinic, space group Pl with lattice constants of a=44.45(2), b=53.11(2), c=32.018(6) Å, [[alpha]]=84.92(2), [[beta]]=95.22(4), [[gamma]]=108.56(4)deg., Z=2, Dobs=1.21gcm-3, Vm=2.07Å3Da-1, Vsol=40.6%. Intensity data were collected using Sakabe camera at BL6A2 of Photon Factory in Tsukuba. The structure of yCyPA was solved by molecular replacement method using hCyPA as a search model (XPLOR program). Refined model of two CyPA molecules and 126 water molecules gave Rfactor of 18.5% for 17931 reflections (F>2[[sigma]]) in the region of 5.0 to 1.9 Å resolution. All C[[alpha]] atoms of two CyPAs are well superimposed with a r.m.s. deviation of 0.23 Å.
yCyPA also has [[beta]]-barrel structure composed of two orthogonal four stranded antiparallel [[beta]]-sheets flanking two [[alpha]]-helixes at the top and bottom as in hCyPA and E. coli CyPA (2). The predicted peptide binding site is identified to be located in the cleft on the upper [[beta]]-sheet. In yCyPA the same residues take the same arrangement as those essential to recognition and binding of peptide-substrates in hCyPA. The hydrophobic pocket, by which peptide-substrates are recognized, is formed by side chains of five non-polar residues; Phe58, Met59, Trpl 19, Leu120 and His124 with
Phe111 at bottom; and Arg53 and Asn100 are positioned on the opposite side of the pocket such that the backbone of the peptide containing a proline can be hydrogen bonded by these two residues to make the proline cis-form.
(1) H. Ke, J. Mol.Biol. (1992) 228, 539-550.
(2) M. Konno et al., J. Mol.Biol. (1996) 256, 897-908.