CGC Bibliography Paper 5482

Structural and biological characterisation of the gut-associated cyclophilin B isoforms from Caenorhabditis elegans.

Picken NC, Eschenlauer S, Taylor P, Page AP, Walkinshaw MD

Medline:

12215411

Citation:

Journal of Molecular Biology 322: 15-25 2002

Type:

ARTICLE

Genes:

cyp-1 cyp-5 cyp-6

Abstract:

The free-living nematode Caenorhabditis elegans expresses 18 cyclophilin isoforms, eight of which are conserved single domain forms, comprising two closely related secreted or type B forms (CYP-5 and CYP-6). Recombinant CYP-5 has been purified, crystallised and the X-ray structure solved to a resolution of 1.75 ANG. The detailed molecular architecture most strongly resembles the structure of human cyclophilin B with conserved changes in loop structure and N and C-terminal extensions. Interestingly, the active site pocket is occupied by a molecule of dithiothreitol though this has little effect on the geometry of the active site which is similar to other cyclophilin structures. The peptidyl-prolyl isomerase activity of CYP-5 has been characterised against the substrate N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide, and gives a kcat/Km value of 3.6X106 M-1 s-1 that compares with a value of 6.3X106 M-1 s-1 for human cyclophilin B. The immunosuppressive drug cyclosporin A binds and inhibits CYP-5 with an IC50 value of 50 nM, which is comparable to the value of 84 nM found for human cyclophilin B. CYP-6 has 67% sequence identity with CYP-5 and a molecular model was built based on the CYP-5 crystal structure. The model shows that CYP-5 and CYP-6 are likely to have very similar structures, but with a markedly increased number of negative charges distributed around the surface of CYP-6. The spatial expression patterns of the cyclophilin B isoforms were examined using transgenic animals carrying a LacZ reporter fusion to these genes, and both cyp-5 and cyp-6 are found to be expressed in an overlapping fashion in the nematode gut. The temporal expression pattern of cyp-5 was further determined and revealed a constitutive