The C-terminal region of human plasma fetuin-B is dispensable for the raised-elephant-trunk mechanism of inhibition of astacin metallopeptidases
2019
Human
fetuin-B plays a key physiological role in
human fertilitythrough its inhibitory action on ovastacin, a member of the
astacinfamily of
metallopeptidases. The inhibitor consists of tandem
cystatin-like domains (CY1 and CY2), which are connected by a linker containing a “CPDCP-trunk” and followed by a C-terminal region (CTR) void of regular secondary structure. Here, we solved the crystal structure of the complex of the inhibitor with archetypal
astacinfrom
crayfish, which is a useful model of human ovastacin. Two hairpins from CY2, the linker, and the tip of the “
legumain-binding loop” of CY1 inhibit
crayfish
astacinfollowing the “raised-
elephant-trunkmechanism” recently reported for mouse
fetuin-B. This inhibition is exerted by blocking active-site cleft sub-sites upstream and downstream of the catalytic zinc ion, but not those flanking the
scissile bond. However, contrary to the mouse complex, which was obtained with
fetuin-B nicked at a single site but otherwise intact, most of the CTR was proteolytically removed during crystallization of the human complex. Moreover, the two complexes present in the crystallographic asymmetric unit diverged in the relative arrangement of CY1 and CY2, while the two complexes found for the mouse complex crystal structure were equivalent. Biochemical studies in vitro confirmed the differential cleavage susceptibility of human and mouse
fetuin-B in front of
crayfish
astacinand revealed that the cleaved human inhibitor blocks
crayfish
astacinand human
meprinα and β only slightly less potently than the intact variant. Therefore, the CTR of animal
fetuin-B orthologs may have a function in maintaining a particular relative orientation of CY1 and CY2 that nonetheless is dispensable for peptidase inhibition.
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