1gi0

File:1gi0.jpg

spinqualitylabelsall models 1gi0, resolution 1.42Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image

A NOVEL SERINE PROTEASE INHIBITION MOTIF INVOLVING A MULTI-CENTERED SHORT HYDROGEN BONDING NETWORK AT THE ACTIVE SITE

OverviewOverview

We describe a new serine protease inhibition motif in which binding is, mediated by a cluster of very short hydrogen bonds (<2.3 A) at the active, site. This protease-inhibitor binding paradigm is observed at high, resolution in a large set of crystal structures of trypsin, thrombin, and, urokinase-type plasminogen activator (uPA) bound with a series of small, molecule inhibitors (2-(2-phenol)indoles and 2-(2-phenol)benzimidazoles)., In each complex there are eight enzyme-inhibitor or enzyme-water-inhibitor, hydrogen bonds at the active site, three of which are very short. These, short hydrogen bonds connect a triangle of oxygen atoms comprising, O(gamma)(Ser195), a water molecule co-bound in the oxyanion hole, (H(2)O(oxy)), and the phenolate oxygen atom of the inhibitor (O6'). Two of, the other hydrogen bonds between the inhibitor and active site of the, trypsin and uPA complexes become short in the thrombin counterparts, extending the three-centered short hydrogen-bonding array into a, tetrahedral array of atoms (three oxygen and one nitrogen) involved in, short hydrogen bonds. In the uPA complexes, the extensive hydrogen-bonding, interactions at the active site prevent the inhibitor S1 amidine from, forming direct hydrogen bonds with Asp189 because the S1 site is deeper in, uPA than in trypsin or thrombin.Ionization equilibria at the active site, associated with inhibitor binding are probed through determination and, comparison of structures over a wide range of pH (3.5 to 11.4) of thrombin, complexes and of trypsin complexes in three different crystal forms. The, high-pH trypsin-inhibitor structures suggest that His57 is protonated at, pH values as high as 9.5. The pH-dependent inhibition of trypsin, thrombin, uPA and factor Xa by 2-(2-phenol)benzimidazole analogs in which, the pK(a) of the phenol group is modulated is shown to be consistent with, a binding process involving ionization of both the inhibitor and the, enzyme. These data further suggest that the pK(a) of His57 of each, protease in the unbound state in solution is about the same, approximately, 6.8. By comparing inhibition constants (K(i) values), inhibitor, solubilities, inhibitor conformational energies and corresponding, structures of short and normal hydrogen bond-mediated complexes, we have, estimated the contribution of the short hydrogen bond networks to, inhibitor affinity ( approximately 1.7 kcal/mol). The structures and K(i), values associated with the short hydrogen-bonding motif are compared with, those corresponding to an alternate, Zn(2+)-mediated inhibition motif at, the active site. Structural differences among apo-enzymes, enzyme-inhibitor and enzyme-inhibitor-Zn(2+) complexes are discussed in, the context of affinity determinants, selectivity development, and, structure-based inhibitor design.

About this StructureAbout this Structure

1GI0 is a Single protein structure of sequence from Bos taurus with CA, MG, SO4 and BMZ as ligands. Active as Trypsin, with EC number 3.4.21.4 Full crystallographic information is available from OCA.

ReferenceReference

A novel serine protease inhibition motif involving a multi-centered short hydrogen bonding network at the active site., Katz BA, Elrod K, Luong C, Rice MJ, Mackman RL, Sprengeler PA, Spencer J, Hataye J, Janc J, Link J, Litvak J, Rai R, Rice K, Sideris S, Verner E, Young W, J Mol Biol. 2001 Apr 13;307(5):1451-86. PMID:11292354

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