Solvation change and ion release during aminoacylation by aminoacyl-tRNA synthetases.
Discrimination between cognate and non-cognate tRNAs by aminoacyl-tRNA synthetases occurs at several steps of the aminoacylation pathway. We have measured changes of solvation and counter-ion distribution at various steps of the aminoacylation pathway of glutamyl- and glutaminyl-tRNA synthetases. The decrease in the association constant with increasing KCl concentration is relatively small for cognate tRNA binding when compared to known DNA-protein interactions. The electro-neutral nature of the tRNA binding domain may be largely responsible for this low ion release stoichiometry, suggesting that a relatively large electrostatic component of the DNA-protein interaction free energy may have evolved for other purposes, such as, target search. Little change in solvation upon tRNA binding is seen. Non-cognate tRNA binding actually increases with increasing KCl concentration indicating that charge repulsion may be a significant component of binding free energy. Thus, electrostatic interactions may have been used to discriminate between cognate and non-cognate tRNAs in the binding step. The catalytic constant of glutaminyl-tRNA synthetase increases with increasing osmotic pressure indicating a water release of 8.4 +/- 1.4 mol/mol in the transition state, whereas little change is seen in the case of glutamyl-tRNA synthetase. We propose that the significant amount of water release in the transition state, in the case of glutaminyl-tRNA synthetase, is due to additional contact of the protein with the tRNA in the transition state.
Banerjee
Rajat
R
Department of Biophysics, Bose Institute, P-1/12, C.I.T. Scheme VII M, Calcutta 700 054, India.
Mandal
Amit Kumar
AK
Saha
Rajesh
R
Guha
Soumi
S
Samaddar
Soma
S
Bhattacharyya
Anusree
A
Roy
Siddhartha
S
D000604:Amino Acyl-tRNA Synthetases / Q000737:chemistry* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005975:Glutamate-tRNA Ligase / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D011189:Potassium Chloride / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D012346:RNA, Transfer, Amino Acyl / Q000737:chemistry* / Q000378:metabolism; D012352:RNA, Transfer, Glu / Q000737:chemistry* / Q000378:metabolism; D012997:Solvents / Q000737:chemistry; D014867:Water / Q000737:chemistry
D012346:RNA, Transfer, Amino Acyl; D012352:RNA, Transfer, Glu; D012997:Solvents; D014867:Water; D011189:Potassium Chloride; D000604:Amino Acyl-tRNA Synthetases; D005975:Glutamate-tRNA Ligase