Electrostatic fingerprints of catalytically active amino acids in enzymes

Abstract

The computed electrostatic and proton transfer properties are studied for 20 enzymes that represent all six major enzyme commission classes and a variety of different folds. The properties of aspartate, glutamate, and lysine residues that have been previously experimentally determined to be catalytically active are reported. The catalytic aspartate and glutamate residues studied here are strongly coupled to at least one other aspartate or glutamate residue and often to multiple other carboxylate residues with intrinsic pK a differences less than 1 pH unit. Sometimes these catalytic acidic residues are also coupled to a histidine residue, such that the intrinsic pK a of the acidic residue is higher than that of the histidine. All catalytic lysine residues studied here are strongly coupled to tyrosine or cysteine residues, wherein the intrinsic pK a of the anion-forming residue is higher than that of the lysine. Some catalytic lysines are also coupled to other lysines with intrinsic pK a differences within 1 pH unit. Some evidence of the possible types of interactions that facilitate nucleophilicity is discussed. The interactions reported here provide important clues about how side chain functional groups that are weak Brønsted acids or bases for the free amino acid in solution can achieve catalytic potency and become strong acids, bases or nucleophiles in the enzymatic environment.

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“…This is achieved through coupling to three nearby histidine residues, the dyad partner H41, the S1 residue H163, and H164, as shown in Table 3 . The strong electrostatic coupling between C145 and these three histidines, with the intrinsic p K a of the anion-forming residue higher than that of each of the cation-forming residues, leads to an expanded buffer range and significant populations of both protonation states that is necessary for catalysis ( Koumanov et al, 2002 ; Ringe et al, 2004 ; Coulther et al, 2021 ; Iyengar et al, 2022 ). Indeed, significant population of the deprotonated state of C145 has been implied in a recent report suggesting covalent binding of selected ligands by C145 ( Mohapatra et al, 2021 ).…”

“…Similarly, the catalytic H41 must have significant population of both protonation states to exchange a proton with the thioester intermediate. This is achieved through the coupling to C145, and also to H164, wherein the two like-charged histidine side chains are strongly coupled to each other and have matched (<1 pH unit difference) intrinsic p K a s ( Table 3 ) ( Koumanov et al, 2002 ; Coulther et al, 2021 ; Iyengar et al, 2022 ).…”

“…Lysine-6968 serves as the base that enables the 2’ oxygen atom of the RNA to attack the methyl group of SAM ( Nencka et al, 2022 ). The deprotonated state of K6968 is significantly populated, first by the strong electrostatic coupling to K6844, wherein the two lysine residues have closely matched intrinsic p K a s ( Table 4 ), and second by strong coupling to Y6845 (and to a lesser extent to Y6930), wherein the anion-forming tyrosine has an intrinsic p K a higher than that of the lysine ( Koumanov et al, 2002 ; Coulther et al, 2021 ; Iyengar et al, 2022 ). Strong coupling to the two anion-forming residues, D6928 and E7001, strengthens the basicity of K6968.…”

“…Pairwise Coulomb potential energies of interaction between amino acid side chains, and the free energies of desolvation of the amino acid side chains, were calculated by a linear Poisson-Boltzmann method ( Antosiewicz et al, 1994 ; Baker et al, 2001 ; Shen et al, 2003 ; Dolinsky et al, 2004 ). The intrinsic p K a s were calculated from the desolvation energy as ( Ullmann, 2003 ; Coulther et al, 2021 ; Iyengar et al, 2022 ): where the intrinsic p K a is the p K a of an amino acid side chain in the hypothetical protein structure where all other ionizable groups are in their electrically neutral state. The model p K a is the p K a of the side chain of the free amino acid in solution.…”

“…Pairwise Coulomb potential energies of interaction between amino acid side chains, and the free energies of desolvation of the amino acid side chains, were calculated by a linear Poisson-Boltzmann method (Antosiewicz et al, 1994;Baker et al, 2001;Shen et al, 2003;Dolinsky et al, 2004). The intrinsic pK a s were calculated from the desolvation energy as (Ullmann, 2003;Coulther et al, 2021;Iyengar et al, 2022):…”