Conservation and function of a potential substrate-binding domain in the yeast Clb5 B-type cyclin

Cyclin A contains a region implicated in binding to the p27 inhibitor and t o substrates. There is strong evolutionary conservation of surface residues contributing to this region in many cyclins, including yeast B-type cyclin s, despite the absence of a yeast p27 homolog. The yeast S-phase B-type cyc lin Clb5p interacted with mammalian p27 in a two-hybrid assay. This interac tion was disrupted by mutations designed to disrupt hydrophobic interaction s (hpm mutation) or hydrogen bonding (Q241A mutation) based on the cyclin A -p27 crystal structure. In contrast, mutation of the Clb5p p27-binding doma in only slightly reduced binding and inhibition by the Sic1p Clb-Cdc28p kin ase inhibitor. Mutations disrupting the p27-binding domain strongly reduced Clb5p biological activity in diverse assays without reducing Clb5p-associa ted kinase activity. An analogous hpm mutation in the mitotic cyclin Clb2p reduced mitotic function, but in some assays this mutation increased the ab ility of Clb2p to perform functions normally restricted to Clb5p. These res ults support the idea of a modular, structurally conserved cyclin domain in volved in substrate targeting.