Tyrosyl tRNA synthetase

Figure 4.15 Schematic diagram of the enzyme tyrosyl-tRNA synthetase, which couples tyrosine to its cognate transfer RNA. The central region of the catalytic domain (red and green) is an open twisted a/p stmcture with five parallel p strands. The active site is formed by the loops from the carboxy ends of P strands 2 and S. These two adjacent strands are connected to a helices on opposite sides of the P sheet.

Figure 4.16 A schematic view of the active site of tyrosyl-tRNA synthetase. Tyrosyl adenylate, the product of the first reaction catalyzed by the enzyme, is bound to two loop regions residues 38-47, which form the loop after p strand 2, and residues 190-193, which form the loop after P strand 5. The tyrosine and adenylate moieties are bound on opposite sides of the P sheet outside the catboxy ends of P strands 2 and 5.

Figure 4.17 Schematic diagram of bound tyrosine to tyrosyl-tRNA synthetase. Colored regions correspond to van der Waals radii of atoms within a layer of the structure through the tyrosine ring. Red is bound tyrosine green is the end of P strand 2 and the beginning of the following loop region yellow is the loop region 189-192 and brown is part of the a helix in loop region 173-177.

Figure 4.18 Side chains of the tyrosyl-tRNA synthetase that form hydrogen bonds to tyrosyl adenylate. Green residues are from p strand 2 and the following loop regions, yellow residues are from the loop after P strand S, and brown residues are from the a helix before P strand S. (Adapted from T. Wells and A. Fersht, Nature 316 656-657, 1985.)...

Brick, R, Bhat, T.N., Blow, D.M. Structure of tyrosyl-tRNA synthetase refined at 2.3 A resolution. Interaction of the enzyme with the tyrosyl adenylate intermediate. /. Mol. Biol. 208 83-98, 1988. 

Scheme 18 Possible carboxylate group participation in the activation of tyrosine in tyrosyl-tRNA synthetase...

Table 7.4 Effects of mutations of Thr40 and His45 on the kinetic parameters for tyrosyl-tRNA synthetase...

Classic doubly wound /3 sheets Lactate dehydrogenase domain 1 Alcohol dehydrogenase domain 2 Aspartate transcarbamylase catalytic domain 2 Phosphoglycerate kinase domain 1 Tyrosyl-tRNA synthetase domain 1( )... 

Tyrosyl-tRNA synthetase dl, d2 Thermolysin dl, d2 T4 phage lysozyme dl, d2 Glucosephosphate isomerase dl, d2 Pyruvate kinase dl, d2 Pyruvate kinase d2, d3 Lactate dehydrogenase dl, d2 Alcohol dehydrogenase dl, d2 Glyceraldehyde-phosphate dehydrogenase dl,d2... 

Chemical reactivity and hydrogen bonding 320 Proton-transfer behaviour 321 Intramolecular hydrogen-bond catalysis 344 Enzyme catalysis and hydrogen bonding 354 Chymotrypsin 354 Thermolysin 355 Carboxypeptidase 355 Tyrosyl tRNA synthetase 356 Summary 366 Acknowledgements 367 References 367... 

The technique of site-directed mutagenesis has been successful in providing details of the mechanism of catalysis by tyrosyl tRNA synthetase. Quantita-... 

Table 17 Effects of hydrogen bonds on free energy differences as measured by AG.pp in tyrosyl activation catalysed by mutations of tyrosyl tRNA synthetase (44).

Fig. 17 Free energy/reaction coordinate diagram for tyrosine activation [see (49)], with wild-type tyrosyl-tRNA synthetase (E) and the Tyr-34 to Phe mutant (E ).

The tyrosyl-tRNA synthetase alters the equilibrium constant for the formation of tyrosyl-adenylate by a factor of 107 by a strain mechanism10 (Chapter 15, section I). 

Figure 4.7 Binding of tyrosine to the tyrosyl-tRNA synthetase from Bacillus stearothermophilus. [From A. R. Fersht, R. S. Mulvey, and G. L. E. Koch, Biochemistry 14,13 (1975).]...

There is no doubt that the enzyme-bound aminoacyl adenylate is formed in the absence of tRNA. It may be isolated by chromatography and the free aminoacyl adenylate obtained by precipitation of the enzyme with acid.47 48 Furthermore, the isolated complex will transfer its amino acid to tRNA. The crystal structure of the tyrosyl-tRNA synthetase bound to tyrosyl adenylate has been solved (Chapter 15, section B). 

Not all aminoacyl-tRNA synthetases have editing sites. The cysteinyl- and tyrosyl-tRNA synthetases bind the correct substrates so much more tightly than their competitors that they do not need to edit.13,14 Similarly, since the accuracy of transcription of DNA by RNA polymerase is better than the overall observed error rate in protein synthesis at about 1 part in 104, RNA polymerases do not need to edit.15 The same should be true for codon-anticodon interactions on the ribosome. However, it is possible that accuracy has been sacrificed to achieve higher rates in this case, which is analogous to a change from Michaelis-Menten to Briggs-Haldane kinetics, and so an editing step is required.16... 

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