The crystal structure of a 250-kDa heterotetrameric particle explains inhibition of sheddase meprin β by endogenous fetuin-B

Abstract

Meprin β (Mβ) is a multidomain type-I membrane metallopeptidase that sheds membrane-anchored substrates, releasing their soluble forms. Fetuin-B (FB) is its only known endogenous protein inhibitor. Herein, we analyzed the interaction between the ectodomain of Mβ (MβΔC) and FB, which stabilizes the enzyme and inhibits it with subnanomolar affinity. The MβΔC:FB crystal structure reveals a ∼250-kDa, ∼160-Å polyglycosylated heterotetrameric particle with a remarkable glycan structure. Two FB moieties insert like wedges through a “CPDCP trunk” and two hairpins into the respective peptidase catalytic domains, blocking the catalytic zinc ions through an “aspartate switch” mechanism. Uniquely, the active site clefts are obstructed from subsites S4 to S10′, but S1 and S1′ are spared, which prevents cleavage. Modeling of full-length Mβ reveals an EGF-like domain between MβΔC and the transmembrane segment that likely serves as a hinge to transit between membrane-distal and membrane-proximal conformations for inhibition and catalysis, respectively.

Helical ultrastructure of the oncogenic metalloprotease meprin α in complex with a small molecule hydroxamate inhibitor

“…However, we were unable to detect any significant differences between meprin α oligomeric forms when assessing small molecule and globular substrates, nor small molecule and globular inhibitors. Indeed, structural studies of human fetuin-B in complex with meprin α are consistent with crystal structures of murine fetuin-B and meprin β (96). Unlike meprin β, these inhibitory complexes were observed to form supramolecular helical packing as a result of the intercalated arrangement of meprin α and numerous fetuin-B molecules.…”

“…Models of the active form and inhibitorbound form were generated similarly, using the model of pro-meprin α as a starting template. Lastly, molecular docking analysis was carried out using the fetuin-B/meprin β crystal structure (PDB-7AUW) (96) superimposed onto the meprin α helix. Subsequently, a model of the meprin α/fetuin-B complex was generated from the model of active meprin α and the AlphaFold model of human fetuin-B (97, 98) (AF-Q9UGM5 v2).…”

A unique network of attack, defence and competence on the outer membrane of the periodontitis pathogen Tannerella forsythia

“…Overall, PotA inhibits MMPs using a “bilobal mechanism” (Fig. 7A and C), which is distantly reminiscent of the “raised elephant trunk mechanism” described for the specific inhibition of astacin MPs by fetuin-B 63,64 and contains elements of the “aspartate-switch mechanism” of latency found in certain MP zymogens. 65 Finally, an OB-fold β-barrel scaffold has also been found in the otherwise unrelated structures of TIMPs.…”

“…Remarkably, the S 1 sub-site of the cle is free, which explains why the inhibitor is not cleaved. In addition, the carboxylate of D 64 provides a strong warhead that binds and thus blocks the catalytic zinc, and further contacts the general base/acid glutamate of MMP-12 (E 219 ; peptidase numbering in subscript, here according to UP P39900; Fig. 7C) or karilysin (E 156 ; for UP, see Fig.…”

Substrate profiling of the metalloproteinase ovastacin – Implications for its physiological function in mammalian fertilization

“…Recently, we were able to elucidate the precise mechanism of this inhibition (22)(23)(24). In mammals, only three members of the astacin family of proteinases, ovastacin and the closely related meprins (α & β) are inhibited by fetuin-B (22,25).…”