Crystal structure of 6‐guanidinohexanoyl trypsin near the optimum pH reveals the acyl‐enzyme intermediate to be deacylated

Abstract

The force driving the conversion from the acyl intermediate to the tetrahedral intermediate in the deacylation reaction of serine proteases remains unclear. The crystal structure of 6-guanidinohexanoyl trypsin was determined at pH 7.0, near the optimum reaction pH, at 1.94 Å resolution. In this structure, three water molecules are observed around the catalytic site. One acts as a nucleophile to attack the acyl carbonyl carbon while the other two waters fix the position of the catalytic water through a hydrogen bond. When the acyl carbonyl oxygen oscillates thermally, the water assumes an appropriate angle to catalyze the deacylation.

Connecting Classical QSAR and LERE Analyses Using Modern Molecular Calculations, LERE‐QSAR (VI): Hydrolysis of Substituted Hippuric Acid Phenyl Esters by Trypsin

“…Further, we confirmed the proposed pose is basically similar to that for the same ligands and papain in our previous docking study (R 1 is also accommodated in the S 1 pocket of papain) 33. The protonation states of titratable residues (Arg, Lys, Asp, Glu, and His) were determined according to their calculated p K a values at pH 8.0, the optimum for the trypsin activity34 by using the PDB2PQR server 36. We treated the protonation states of the side‐chains of His57, Asp102, and Ser195 in the catalytic triad as neutral (N δ H), anionic, and neutral (O γ H) forms, respectively.…”

Polymorphic Transformation of Antibiotic Clarithromycin Under Acidic Condition

“…The rate constants increase as pH decreases. As the solvent content of form A is thought to be very high as described above, chloride ions might not be tightly fixed in the form A crystal and, moreover, might be freely exchanged with those in the bulk solution outside the crystal, as is usual in macromolecule crystal of high solvent content . The chloride ion in form A may be in equilibrium between ionically bonded state to the dimethyl aminomethyl moiety of CAM and freely diffusing state in the large solvent region in the crystal.…”

Linear Discriminant Analysis for the <i>in Silico</i> Discovery of Mechanism-Based Reversible Covalent Inhibitors of a Serine Protease: Application of Hydration Thermodynamics Analysis and Semi-empirical Molecular Orbital Calculation

“…Preparation of Ligand Datasets The covalent ligands [27][28][29][30][31][32][33][34] were separated into two groups-inhibitor group and substrate group, according to their Log(k cat ) at a cut-off of −1.0. The cut-off was set because it allowed clear differentiation between the inhibitor group-including clinically adopted drugs, nafamostat (Log(k cat )=−4.5) and gabexate (Log(k cat )=−3.1)and the substrate group mainly consisting of substrate-analog compounds [e.g., K3 (Log(k cat )=−0.11)] and other substrateanalog peptides [e.g., succinyl-alanyl-alanyl-prolyl-lysyl (suc-AAPK) (Log(k cat )=1.6)].…”

“…In those ligands, nafamostat has two corresponding crystal structures (1GBT and 2AH4). Acyltrypsin intermediates of the other ligands with no corresponding crystal structures 27,28,30) were modeled by flexible molecular superposition using Phase 22,23,39) (Schrödinger suite 2015-3). Prolyl-arginyl-catalytic Ser195 moiety in the crystal structure of suc-AAPR trypsin (2AGE) 31) was used as the reference structure of covalent ligands in the acyl-trypsin intermediate.…”