Hepatocyte growth factor activator (HGFA): molecular structure and interactions with HGFA inhibitor-1 (HAI-1).
The trypsin-like serine protease hepatocyte growth factor activator (HGFA) undergoes proteolytic activation during blood coagulation, resulting in a 34 kDa 'short form', consisting mainly of the protease domain. The crystal structures of the recombinantly expressed HGFA 'short form' discussed herein have provided molecular insights into its interaction with inhibitors and substrates, as well as the regulation of catalytic activity. The HGFA structures revealed enzymatically competent and noncompetent forms associated with the conformational states of two substrate specificity-determining loops, the 220-loop and 99-loop. The implied dynamic behavior of these loops, which are intimately involved in substrate interaction, has precedents in other members of the S1 family of serine proteases, and may be associated with specific mechanisms of enzyme regulation. Furthermore, HGFA activity is strongly inhibited by HGFA inhibitor-1, a membrane-spanning multidomain inhibitor containing two Kunitz domains, of which only the N-terminal Kunitz domain-1 (KD1) inhibits enzymatic activity. In the structure of the KD1-HGFA complex, the inhibitor interacts with the active site region by making contacts with all substrate specificity-determining loops and by occupying subsites S1, S2 and S4 in a substrate-like manner. In fact, the side chains of KD1 residues occupying these sites are virtually superimposable on the P1, P2 and P4 residues of the pro-hepatocyte growth factor-derived substrate mimic Lys-Gln-Leu-Arg chloromethyl ketone bound to HGFA. These structures also allow us to rationalize the apparently narrow substrate specificity of HGFA, which is limited to the two known macromolecular substrates pro-hepatocyte growth factor and pro-macrophage-stimulating protein.
Eigenbrot
Charles
C
Department of Structural Biology, Genentech, Inc., South San Francisco, CA 94080, USA.
Ganesan
Rajkumar
R
Kirchhofer
Daniel
D
D000818:Animals; D020134:Catalytic Domain; D006801:Humans; D011487:Protein Conformation; D053491:Proteinase Inhibitory Proteins, Secretory / Q000737:chemistry* / Q000378:metabolism*; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000378:metabolism*
D053491:Proteinase Inhibitory Proteins, Secretory; C109149:SPINT1 protein, human; C081092:HGF activator; D012697:Serine Endopeptidases