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Engineering an Mg 2+ site to replace a structurally conserved arginine in the catalytic center of histidyl‐tRNA synthetase by computer experiments
Author(s) -
Arnez John G.,
Flanagan Karen,
Moras Dino,
Simonson Thomas
Publication year - 1998
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/(sici)1097-0134(19980815)32:3<362::aid-prot11>3.0.co;2-7
Subject(s) - transfer rna , arginine , center (category theory) , catalysis , chemistry , biochemistry , stereochemistry , computational biology , amino acid , biology , rna , crystallography , gene
Histidyl‐tRNA synthetase (HisRS) differs from other class II aminoacyl‐tRNA synthetases (aaRS) in that it harbors an arginine at a position where the others bind a catalytic Mg 2+ ion. In computer experiments, four mutants of HisRS from Escherichia coli were engineered by removing the arginine and introducing a Mg 2+ ion and residues from seryl‐tRNA synthetase (SerRS) that are involved in Mg 2+ binding. The mutants recreate an active site carboxylate pair conserved in other class II aaRSs, in two possible orders: Glu‐Asp or Asp‐Glu, replacing Glu‐Thr in native HisRS. The mutants were simulated by molecular dynamics in complex with histidyl‐adenylate. As controls, the native HisRS was simulated in complexes with histidine, histidyl‐adenylate, and histidinol. The native structures sampled were in good agreement with experimental structures and biochemical data. The two mutants with the Glu‐Asp sequence showed significant differences in active site structure and Mg 2+ coordination from SerRS. The others were more similar to SerRS, and one of them was analyzed further through simulations in complex with histidine, and His+ATP. The latter complex sampled two Mg 2+ positions, depending on the conformation of a loop anchoring the second carboxylate. The lowest energy conformation led to an active site geometry very similar to SerRS, with the principal Mg 2+ bridging the α‐ and β‐phosphates, the first carboxylate (Asp) coordinating the ion through a water molecule, and the second (Glu) coordinating it directly. This mutant is expected to be catalytically active and suggests a basis for the previously unexplained conservation of the active site Asp‐Glu pair in class II aaRSs other than HisRS. Proteins 32:362–380, 1998. © 1998 Wiley‐Liss, Inc.