LYSYL-TRANSFER RNA-SYNTHETASE FROM YEAST - DISCRIMINATION OF AMINO-ACIDS BY NATIVE AND PHOSPHORYLATED SPECIES

被引:13
作者
FREIST, W
STERNBACH, H
CRAMER, F
机构
[1] Max-Planck-Institut Für Experimentelle Medizin, Gottingen
来源
EUROPEAN JOURNAL OF BIOCHEMISTRY | 1992年 / 204卷 / 03期
关键词
D O I
10.1111/j.1432-1033.1992.tb16723.x
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Discrimination factors (D) which are characteristic for discrimination between lysine and 19 naturally occurring Don-cognate amino acids have been determined from k(cat) and K(m) values for native and phosphorylated lysyl-tRNA synthetases from yeast. Generally, both species of this class II aminoacyl-tRNA synthetase are considerably less specific than the class I synthetases specific for isoleucine, valine, tyrosine, and arginine. D values of the native enzyme are in the range 90-1700, D values of the phosphorylated species in the range 40-770. The phosphorylated enzyme acts faster and less accurately. In aminoacylation of tRNA(Lys)-C-C-A(2'NH2) discrimination factors D1 vary over 30-980 for the native and over 8-300 for the phosphorylated enzyme. From AMP formation stoichiometry and D1 values pretransfer proof-reading factors (PI-1) of 1.1-56 were calculated for the native enzyme, factors of 1.0-44 for the phosphorylated species. Post-transfer proof-reading factors (PI-2) were calculated from D values and AMP formation stoichiometry in acylation of tRNA(Lys)-C-C-A. Pretransfer proof-reading is the main correction step, posttransfer proof-reading is less effective or negligible (PI-2 almost-equal-to 1-8). Initial discrimination factors (I), which are due to differences in Gibbs free energies of binding between lysine and noncognate substrates (DELTA-DELTA-G(I)), were calculated from discrimination and proofreading factors. In contrast to class I synthetases, for lysyl-tRNA synthetase only one initial discrimination step can be assumed and amino acid recognition is reduced to a three-step process instead of the four-step recognition observed for the class I synthetases. Plots of DELTA-DELTA-G(I) values against accessible surface areas of amino acids show clearly that phosphorylation of the enzyme changes the structures of the amino acid binding sites. This is illustrated by a hypothetical 'stopper model' of these sites.
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页码:1015 / 1023
页数:9
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