Molecular Mechanism of Amino Acid Discrimination for the Editing Reaction of E.coli Leucyl-tRNA Synthetase

Authors: Keun Woo Lee, Minky Son, Chanin Park, Ayoung Baek

Abstract:

Certain tRNA synthetases have developed highly accurate molecular machinery to discriminate their cognate amino acids. Those aaRSs achieve their goal via editing reaction in the Connective Polypeptide 1 (CP1). Recently mutagenesis studies have revealed the critical importance of residues in the CP1 domain for editing activity and X-ray structures have shown binding mode of noncognate amino acids in the editing domain. To pursue molecular mechanism for amino acid discrimination, molecular modeling studies were performed. Our results suggest that aaRS bind the noncognate amino acid more tightly than the cognate one. Finally, by comparing binding conformations of the amino acids in three systems, the amino acid binding mode was elucidated and a discrimination mechanism proposed. The results strongly reveal that the conserved threonines are responsible for amino acid discrimination. This is achieved through side chain interactions between T252 and T247/T248 as well as between those threonines and the incoming amino acids.

Keywords: Amino acid discrimination, Binding free energy Leucyl-tRNAsynthetase, Molecular dynamics.

Digital Object Identifier (DOI): doi.org/10.5281/zenodo.1056366

References:

[1] Carter Jr., C. W.,"Cognition, mechanism, and evolutionary relationships in aminoacyl-tRNAsynthetases",Annu. Rev. Biochem,vol. 62, pp. 715-748, 1993.
[2] Martinis, S.A., and Schimmel, P.,"Escherichia coli and Salmonella Cellular and Molecular Biology", 2nd Ed., ASM, Neidhardt FC, Washington DC, 1996.
[3] Giege, R., Sissler, M., Florentz, C.,"Universal rules and idiosyncratic features in tRNA identity",Nucleic Acids Res,vol. 26, pp. 5017-5035,1998.
[4] Fersht, A. R., Kaethner, M. M., "Mechanism of aminoacylation of tRNA. Proof of the aminoacyladenylate pathway for the isoleucyl- and tyrosyl-tRNAsynthetases from Escherichia coli K12", Biochemistry,vol. 15, pp. 818-823,1976.
[5] Pauling, L., Festschrift f├╝r Prof. Dr. Arthur Stoll, BirkhauserVerlag, Basel 1958.
[6] Loftfield, R. B.,"The Frequency of errors in protein biosynthesis",Biochem. J.,vol. 89, pp. 82-92, 1963.
[7] Eldred, E. W., Schimmel, P.,"Rapid deacylation by isoleucyl transfer ribonucleic acid synthetase of isoleucine-specific transfer ribonucleic acid aminoacylated with valine", J. Biol. Chem.,vol. 247, pp. 2961-2964, 1972.
[8] Schmidt, E., Schimmel, P.,"Insights into editing from an ile-tRNAsynthetase structure with tRNAile and mupirocin",Science,vol. 285, pp. 1074-1077, 1994.
[9] Hale, S. P., Schimmel, P.,"Protein synthesis editing by a DNA aptamer",Proc. Natl. Acad. Sci. USA,vol. 93, pp. 2755-2758, 1996.
[10] Fersht, A. R., Kaethner, M. M.,"Enzyme hyperspecificity. Rejection of threonine by the valyltRNAsynthetase by misacylation and hydrolytic editing",Biochemistry,vol. 15, pp. 3342-3346, 1976.
[11] Fersht, A. R.,"Editing mechanisms in protein synthesis. Rejection of valine by the isoleucyl-tRNAsynthetase",Biochemistry,vol. 16, pp. 1025-1030, 1977.
[12] Fersht, A. R.,"Enzyme Structure and Mechanism", Freeman, San Francisco, 1997.
[13] Fersht, A. R., Dingwall C.,"Establishing the misacylation/deacylation of the tRNA pathway for the editing mechanism of prokaryotic and eukaryotic valyl-tRNAsynthetases",Biochemistry,vol. 18, pp. 1238-1245, 1979.
[14] Fersht, A. R.,"Sieves in sequence",Science,vol. 280, pp. 541, 1998.
[15] Nureki, O., Vassylyev D. G., Tateno, M., Shimada, A., Nakama, T., Fukai, S., Konno, M., Hendrickson, T.L., Schimmel, P., Yokoyama, S.,"Enzyme structure with two catalytic sites for double-sieve selection of substrate",Science,vol. 280, pp. 578-582, 1998.
[16] Silvian, L. F., Wang, J., Steitz, T. A.,"Insights into editing from an ile-tRNAsynthetase structure with tRNAile and mupirocin",Science,vol. 285, pp. 1074-1077, 1999.
[17] Fukai, S., Nureki, O., Sekine, S., Shimada, A., Tao, J., Vassylyev, D. G., Yokoyama, S.,"Structural basis for double-sieve discrimination of L-Val from L-Ile and L-threonine by the complex of tRNAVal and valyl-tRNAsynthetase",Cell,vol. 103, pp. 793-893, 2000.
[18] Cusack, S., Yaremchuk, A., Tukalo, M.,"The 2 Å crystal structure of leucyl-tRNAsynthetase and its complex with a leucyl-adenylate analogue",EMBO J.,vol. 19, pp. 2351-2631, 2000.
[19] Lincecum Jr., T. L., Tukalo, M,, Yaremchuk, A., Mursinna, R. S., Williams, A. M., Sproat, B. S., Van Den Eynde, W., Link, A., Van Calenbergh, S., Grotli, M., Martinis, S. A., Cusack, S.,"Structural and mechanistic basis of pre- and posttransfer editing by leucyl-tRNAsynthetase",Mol. Cell,vol. 11, pp. 951-963, 2003.
[20] Bishop, A. C., Nomanbhoy, T. K., Schimmel, P., "locking site-to-site translocation of a misactivated amino acid by mutation of a class I tRNAsynthetase", Proc. Natl. Acad. Sci. USA,vol. 99, pp. 585-590, 2002.
[21] Hendrickson, T. L., Nomanbhoy, T. K., Schimmel, P.,"Errors from selective disruption of the editing center in a tRNAsynthetase",Biochemistry,vol. 39, pp. 8180-8186,2000.
[22] Mursinna, R. S., Lincecum, T. L., Martinis, S. A.,"A conserved threonine within Escherichia coli leucyl-tRNAsynthetase prevents hydrolytic editing of leucyl-tRNALeu",Biochemistry,vol. 40, pp. 5376-5381, 2001.
[23] Mursinna, R. S., Martinis, S. A., "Rational design to block amino acid editing of a tRNAsynthetase",J. Am. Chem. Soc.,vol. 124, pp. 7286-7287, 2002.
[24] Mursinna, R. S., Lee, K. W., Briggs, J. M., Martinis, S. A.,"Molecular dissection of a critical specificity determinant within the amino acid editing domain of leucyl-tRNAsynthetase",Biochemistry,vol. 43, pp. 155-165, 2004.
[25] Lee, K. W., Briggs, J. M.,"Molecular modeling study of the editing active site of Escherichia coli leucyl-tRNAsynthetase: two amino acid binding sites in the editing domain",Proteins: Struct. Funct. Bioinformatics,vol. 54,pp. 693-704, 2004.
[26] Brooks, B. R., Bruccoleri, R. E., Olafson, B. D., States, D. J., Swaminathan, S., Karplus, M.,"CHARMM: A program for macromolecular energy, minimisation and dynamics calculations",J. Comp. Chem.,vol. 4, pp. 187-217, 1983.
[27] Kale, L., Skeel, R., Bhandarkar, M., Brunner, R., Gursoy, A., Krawetz, N., Phillips, J., Shinozaki, A., Varadarajan, K., Schulten, K.,"NAMD2: Greater Scalability for Parallel Molecular Dynamics",J. Comp. Phys.,vol. 151, pp. 283-312,1999.
[28] Berendsen, H. J. C., Postma, J. P. M., van Gunsteren, W. F., DiNola, A., Haak, J. R.,"Molecular dynamics with coupling to an external bath",J. Chem. Phys.,vol. 81, pp. 3684-3690,1984.
[29] Amadei, A., Linssen, A. B. M., Berendsen, H. J. C., "Essential dynamics of proteins",Proteins: Struct. Funct. Genetics,vol. 17, pp. 412-425,1993.
[30] Vriend, G.,"WHAT IF: A molecular modeling and drug design program",J. Mol. Graph.,vol. 8, pp. 52-56, 1990.
[31] Aqvist, J., Medina, C., Samuel, J. E.,"A new method for predicting binding affinity in computer-aided drug design",Protein Eng.,vol. 7, pp. 385-391, 1994.
[32] Hansson, T., Åqvist, J.,"Estimation of binding free energies for HIV proteinase inhibitors by molecular dynamics simulations",Protein Eng.,vol. 8, pp. 1137-1144,1995.
[33] Aqvist, J.,"Calculation of absolute binding free energies for charged ligands and effects of long-range electrostatic interactions",J. Comp. Chem.,vol. 17, pp. 1587-1597,1996.
[34] Srinivasan, J., Cheatham, T. E., Cieplak, P., Kollman, P. A., Case, D. A.,"Continuum Solvent Studies of the Stability of DNA, RNA, and Phosphoramidate-DNA Helices",J. Am. Chem. Soc.,vol. 120, pp. 9401-9409, 1998.
[35] Kollman, P. A., Massova, I., Reyes, C., Kuhn, B., Huo, S., Chong, L., Lee, M., Lee, T., Duan, Y., Wang, W., Donini, O., Cieplak, P., Srinivasan, J., Case, D. A., Cheatham, T. E. III.,"Calculating structures and free energies of complex molecules: combining molecular mechanics and continuum models",Acc. Chem. Res.,vol. 33, pp. 889-897, 2000.