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either active site or exosite I affect thrombin specificity among PARs (Ayala et al.
2001
). This behavior of substrates and inhibitors extending from active site and
wrapping around a corner to reach an external site may be relevant to how some
MMPs might grip protein fibrils for cleavage (see Sects.
6.2.2.2
,
6.2.2.3
,
6.2.2.4
,
and
6.3.1
).
Like the hirudin complex, the haemadin complex is a case were the inhibitory
polypeptide reaches from N-terminal interactions with the thrombin active site to its
C-terminal acidic tail interaction with basic exosite. However, the exosite is exosite II
near the “top” of the protease in the standard view (Richardson et al.
2000
). The
binding of heparin by exosite II of thrombin facilitates the binding of antithrombin or
protein C inhibitor by bridging between exosite II and the serpin's positively charged
heparin-binding site, which is especially clear in the ternary complex with antithrom-
bin (Dementiev et al.
2004
;Lietal.
2004
,
2008a
,
b
). The concept of GAG chain
bridging between the positive patch of a protease and the positive patch of a partner
may be relevant to GAG-dependent stimulation both of activation of pro-MMP-7, -1,
and -2 (Sect.
6.1.2
) and of MMP-7 activity upon pro-
-defensins (next section).
a
6.2.2.2 Matrix-Interacting Exosites in Catalytic Domain
Enhance Proteolysis
One early hint for the possibility of allosteric effectors for MMPs was heparin-
increased activity of MMP-7, -1, or -13 in zymography (Yu and Woessner
2001
).
While the activation mechanism remains unclear, in the case of MMP-7, associa-
tion with heparin around the distal positively charged cradle of residues is likely
(Fig.
6.1
). The GAG-dependent activation of MMP-7 in the maturation of pro-
a
-defensins (Ra et al.
2009
) has the additional mechanistic possibility of simply
bringing the protease and substrate together as described above. The importance of
the V-B loop in collagenolysis by MMP-1 and -8 (Chung et al.
2000
; Pelman et al.
2005
) (at right in Fig.
6.2
) outside the conserved heart of the active site cleft is
consistent with an exosite in that region, as described below. Evidence of peptide
substrates “turning a corner” to interact with this region is implied by the influence
of the P4
0
and even P5
0
and P6
0
positions in MMP-2 cleavage of peptides from
libraries (Schilling and Overall
2008
).
6.2.2.3 Elastin Interactions Beyond the Active Site
The concept of an exosite from thrombin research was recently established both in
structural and functional terms within the catalytic domain of MMP-12. The catalytic
domain of MMP-12 is the predominant, mature form observed in vivo, making it
like MMP-7 in practice. Much of the active site cleft where elastin peptides were
computationally docked with MMP-12 (Bertini et al.
2009
) is highly conserved.
That conservation raises the question of what determines MMP-12's relatively high
specific activity toward elastin? At least one
exosite
and ten residues of five loops
