Limits...
Catalytic mechanism of the tryptophan activation reaction revealed by crystal structures of human tryptophanyl-tRNA synthetase in different enzymatic states.

Shen N, Zhou M, Yang B, Yu Y, Dong X, Ding J - Nucleic Acids Res. (2008)

Bottom Line: The dimeric hTrpRS is structurally and functionally asymmetric with half-of-the-sites reactivity.The KMSAS loop appears to have an inherent flexibility and the binding of ATP stabilizes it in a closed conformation that secures the position of ATP for catalysis.Our structural data indicate that the catalytic mechanism of the Trp activation reaction by hTrpRS involves more moderate conformational changes of the structural elements at the active site to recognize and bind the substrates, which is more complex and fine-tuned than that of bacterial TrpRS.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, and Graduate School of Chinese Academy of Sciences, 320 Yue-Yang Road, Shanghai 200031, China.

ABSTRACT
Human tryptophanyl-tRNA synthetase (hTrpRS) differs from its bacterial counterpart at several key positions of the catalytic active site and has an extra N-terminal domain, implying possibly a different catalytic mechanism. We report here the crystal structures of hTrpRS in complexes with Trp, tryptophanamide and ATP and tryptophanyl-AMP, respectively, which represent three different enzymatic states of the Trp activation reaction. Analyses of these structures reveal the molecular basis of the mechanisms of the substrate recognition and the activation reaction. The dimeric hTrpRS is structurally and functionally asymmetric with half-of-the-sites reactivity. Recognition of Trp is by an induced-fit mechanism involving conformational change of the AIDQ motif that creates a perfect pocket for the binding and activation of Trp and causes coupled movements of the N-terminal and C-terminal domains. The KMSAS loop appears to have an inherent flexibility and the binding of ATP stabilizes it in a closed conformation that secures the position of ATP for catalysis. Our structural data indicate that the catalytic mechanism of the Trp activation reaction by hTrpRS involves more moderate conformational changes of the structural elements at the active site to recognize and bind the substrates, which is more complex and fine-tuned than that of bacterial TrpRS.

Show MeSH
Conformational differences of the substrate-binding pocket. (A) Electrostatic surfaces of the substrate-binding pocket in different enzymatic states, showing the changes of the size and shape of the pocket. (B) A graph diagram showing the changes of the solvent-accessible surface area and the volume of the pocket. For parallel comparison, the N-terminal β-hairpin in the pretransition and product states is omitted in the surface presentation and the calculation of the solvent-accessible surface area and the volume of the pocket.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2275098&req=5

Figure 3: Conformational differences of the substrate-binding pocket. (A) Electrostatic surfaces of the substrate-binding pocket in different enzymatic states, showing the changes of the size and shape of the pocket. (B) A graph diagram showing the changes of the solvent-accessible surface area and the volume of the pocket. For parallel comparison, the N-terminal β-hairpin in the pretransition and product states is omitted in the surface presentation and the calculation of the solvent-accessible surface area and the volume of the pocket.

Mentions: The structure of the hTrpRS–TrpNH2O–ATP complex is the first structure of hTrpRS bound with ATP and represents the pretransition state of the enzyme. Thus, analysis of this complex allows us to identify the structural elements and residues that are involved in the recognition and binding of ATP and/or play important roles in the aminoacylation reaction. In this complex, both the AIDQ motif and the KMSAS loop assume the closed conformations, forming a closed substrate-binding pocket (Figure 3 and Table 2). Meanwhile, both the N-terminal and C-terminal domains also adopt the closed conformations, assuming the closed overall conformation. The closed conformation of the KMSAS loop is necessary to assemble a proper ATP-binding site to accommodate the substrate and carry out the Trp activation reaction.Figure 3.


Catalytic mechanism of the tryptophan activation reaction revealed by crystal structures of human tryptophanyl-tRNA synthetase in different enzymatic states.

Shen N, Zhou M, Yang B, Yu Y, Dong X, Ding J - Nucleic Acids Res. (2008)

Conformational differences of the substrate-binding pocket. (A) Electrostatic surfaces of the substrate-binding pocket in different enzymatic states, showing the changes of the size and shape of the pocket. (B) A graph diagram showing the changes of the solvent-accessible surface area and the volume of the pocket. For parallel comparison, the N-terminal β-hairpin in the pretransition and product states is omitted in the surface presentation and the calculation of the solvent-accessible surface area and the volume of the pocket.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2275098&req=5

Figure 3: Conformational differences of the substrate-binding pocket. (A) Electrostatic surfaces of the substrate-binding pocket in different enzymatic states, showing the changes of the size and shape of the pocket. (B) A graph diagram showing the changes of the solvent-accessible surface area and the volume of the pocket. For parallel comparison, the N-terminal β-hairpin in the pretransition and product states is omitted in the surface presentation and the calculation of the solvent-accessible surface area and the volume of the pocket.
Mentions: The structure of the hTrpRS–TrpNH2O–ATP complex is the first structure of hTrpRS bound with ATP and represents the pretransition state of the enzyme. Thus, analysis of this complex allows us to identify the structural elements and residues that are involved in the recognition and binding of ATP and/or play important roles in the aminoacylation reaction. In this complex, both the AIDQ motif and the KMSAS loop assume the closed conformations, forming a closed substrate-binding pocket (Figure 3 and Table 2). Meanwhile, both the N-terminal and C-terminal domains also adopt the closed conformations, assuming the closed overall conformation. The closed conformation of the KMSAS loop is necessary to assemble a proper ATP-binding site to accommodate the substrate and carry out the Trp activation reaction.Figure 3.

Bottom Line: The dimeric hTrpRS is structurally and functionally asymmetric with half-of-the-sites reactivity.The KMSAS loop appears to have an inherent flexibility and the binding of ATP stabilizes it in a closed conformation that secures the position of ATP for catalysis.Our structural data indicate that the catalytic mechanism of the Trp activation reaction by hTrpRS involves more moderate conformational changes of the structural elements at the active site to recognize and bind the substrates, which is more complex and fine-tuned than that of bacterial TrpRS.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, and Graduate School of Chinese Academy of Sciences, 320 Yue-Yang Road, Shanghai 200031, China.

ABSTRACT
Human tryptophanyl-tRNA synthetase (hTrpRS) differs from its bacterial counterpart at several key positions of the catalytic active site and has an extra N-terminal domain, implying possibly a different catalytic mechanism. We report here the crystal structures of hTrpRS in complexes with Trp, tryptophanamide and ATP and tryptophanyl-AMP, respectively, which represent three different enzymatic states of the Trp activation reaction. Analyses of these structures reveal the molecular basis of the mechanisms of the substrate recognition and the activation reaction. The dimeric hTrpRS is structurally and functionally asymmetric with half-of-the-sites reactivity. Recognition of Trp is by an induced-fit mechanism involving conformational change of the AIDQ motif that creates a perfect pocket for the binding and activation of Trp and causes coupled movements of the N-terminal and C-terminal domains. The KMSAS loop appears to have an inherent flexibility and the binding of ATP stabilizes it in a closed conformation that secures the position of ATP for catalysis. Our structural data indicate that the catalytic mechanism of the Trp activation reaction by hTrpRS involves more moderate conformational changes of the structural elements at the active site to recognize and bind the substrates, which is more complex and fine-tuned than that of bacterial TrpRS.

Show MeSH