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studies show that there is not sufficient room in the S1'-S3' cleft to accommodate the native coiled triple
collagen bundle.
The active site consists of a series of subsites on either side of the catalytic zinc. These subsites are
numbered starting at the catalytic zinc and preceding from N to C as S1', S2', etc., corresponding to the
residues (P1', P2', etc.) of the substrate that is bound (Figure 2). Likewise the subsites are numbered S1,
S2, etc. outward from the other side of the catalytic zinc.
While the interaction of the substrate (and the inhibitor) with the catalytic zinc is the most important
interaction, the remainder of the substrate (inhibitor) also forms hydrogen bonds with residues from the
top strand of the β sheet and the loop region posterior to the Met-turn. These interactions with the
substrate in the binding pockets of the MMPs are the prime targets for engineering specific MMP
inhibitors. An in-depth understanding of the differences of the properties of these pockets in the different
MMPs and the interactions of specific residues within these pockets is essential for structure-based
design of inhibitors.
IV. Main-Chain Substrate Interactions
Most of the hydrogen bonds between the substrate and the MMP occur with the top strand of the β sheet.
The P3 residue does not make any direct hydrogen bonds with the MMP. The P2 residue makes two
hydrogen bonds with residue 184, which is a conserved alanine residue in all the aligned MMP
sequences. Residue 183 is a conserved histidine, which is bound to the structural zinc, further stabilizing
the conformation of the top strand. The carbonyl oxygen of P1 is liganded to the catalytic zinc. The lefthand
side of the substrate is thus held in place by only two hydrogen bonds with the enzyme and one
interaction with the catalytic zinc. Although the P3 residue does not make any hydrogenbond
contributions to substrate binding, it is essential for catalytic activity [43].
The right-hand side of the substrate is held much tighter. The P1' residue's carbonyl oxygen makes a
hydrogen bond with the amide nitrogen of residue 181, which is a conserved leucine residue. The amide
nitrogen of the P1' residue is hydrogen bonded to the conserved alanine-182 carbonyl oxygen. The P2'
substrate residue is held in place by hydrogen bonds to proline 238 and tyrosine 240, two more
conserved residues. The amide nitrogen of P3' makes a hydrogen bond with the carbonyl oxygen of
residue 179, the only nonconserved residue, making a main-chain interaction with the substrate.
The use of conserved residues to maintain the main-chain interactions along the substrate backbone
makes differentiation of the MMPs through these interactions difficult. Instead, differences in the
regions where the side chains of the substrate interact can be used to drive the discovery of specific
MMP inhibitors. The hydrogen-bonding pattern also indicates that right-hand side (P'-
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