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Identification and analysis of residues contained on β → a loops of the dual-substrate (βα)8 phosphoriblosyl isomerase A specific for its phosphoribosyl anthranilate isomerase activity
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Noda-Garcia, Lianet, Camacho-Zarco, Aldo R., Verdel-Aranda, Karina, Wright, H. (Helena), Soberon, Xavier, Fülöp, Vilmos and Barona-Gomez, Francisco (2010) Identification and analysis of residues contained on β → a loops of the dual-substrate (βα)8 phosphoriblosyl isomerase A specific for its phosphoribosyl anthranilate isomerase activity. Protein Science, Vol.19 (No.3). pp. 535-543. doi:10.1002/pro.331 ISSN 0961-8368.
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Official URL: http://dx.doi.org/10.1002/pro.331
Abstract
A good model to experimentally explore evolutionary hypothesis related to enzyme function is the ancient-like dual-substrate (βα)8 phosphoribosyl isomerase A (PriA), which takes part in both histidine and tryptophan biosynthesis in Streptomyces coelicolor and related organisms. In this study, we determined the Michaelis–Menten enzyme kinetics for both isomerase activities in wild-type PriA from S. coelicolor and in selected single-residue monofunctional mutants, identified after Escherichia coliin vivo complementation experiments. Structural and functional analyses of a hitherto unnoticed residue contained on the functionally important β → α loop 5, namely, Arg139, which was postulated on structural grounds to be important for the dual-substrate specificity of PriA, is presented for the first time. Indeed, enzyme kinetics analyses done on the mutant variants PriA_Ser81Thr and PriA_Arg139Asn showed that these residues, which are contained on β → α loops and in close proximity to the N-terminal phosphate-binding site, are essential solely for the phosphoribosyl anthranilate isomerase activity of PriA. Moreover, analysis of the X-ray crystallographic structure of PriA_Arg139Asn elucidated at 1.95 Å herein strongly implicates the occurrence of conformational changes in this β → α loop as a major structural feature related to the evolution of the dual-substrate specificity of PriA. It is suggested that PriA has evolved by tuning a fine energetic balance that allows the sufficient degree of structural flexibility needed for accommodating two topologically dissimilar substrates—within a bifunctional and thus highly constrained active site—without compromising its structural stability.
Item Type: | Journal Article | ||||
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Subjects: | Q Science > QD Chemistry | ||||
Divisions: | Faculty of Science, Engineering and Medicine > Science > Life Sciences (2010- ) > Biological Sciences ( -2010) | ||||
Journal or Publication Title: | Protein Science | ||||
Publisher: | John Wiley & Sons Ltd. | ||||
ISSN: | 0961-8368 | ||||
Official Date: | March 2010 | ||||
Dates: |
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Volume: | Vol.19 | ||||
Number: | No.3 | ||||
Number of Pages: | 9 | ||||
Page Range: | pp. 535-543 | ||||
DOI: | 10.1002/pro.331 | ||||
Status: | Peer Reviewed | ||||
Publication Status: | Published | ||||
Access rights to Published version: | Restricted or Subscription Access | ||||
Funder: | Royal Society (Great Britain), Conacyt, Mexico, Royal Society, United Kingdom | ||||
Grant number: | 50952-0, 83039 |
Data sourced from Thomson Reuters' Web of Knowledge
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