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Structure of the R65Q Mutant of Yeast 3-Phosphoglycerate Kinase Complexed with Mg-AMP-PNP and 3-Phospho-d-glycerate

McPhillips, Timothy M. and Hsu, Barbara T. and Sherman, Mark A. and Mas, Maria T. and Rees, Douglas C. (1996) Structure of the R65Q Mutant of Yeast 3-Phosphoglycerate Kinase Complexed with Mg-AMP-PNP and 3-Phospho-d-glycerate. Biochemistry, 35 (13). pp. 4118-4127. ISSN 0006-2960. http://resolver.caltech.edu/CaltechAUTHORS:20150205-140310848

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Abstract

The structure of a ternary complex of the R65Q mutant of yeast 3-phosphoglycerate kinase (PGK) with magnesium 5‘-adenylylimidodiphosphate (Mg-AMP-PNP) and 3-phospho-d-glycerate (3-PG) has been determined by X-ray crystallography to 2.4 Å resolution. The structure was solved by single isomorphous replacement, anomalous scattering, and solvent flattening and has been refined to an R-factor of 0.185, with rms deviations from ideal bond distance and angles of 0.009 Å and 1.78°, respectively. PGK consists of two domains, with the 3-PG bound to a “basic patch” of residues from the N-terminal domain and the Mg-AMP-PNP interacting with residues from the C-terminal domain. The two ligands are separated by ∼11 Å across the interdomain cleft. The model of the R65Q mutant of yeast PGK is very similar to the structures of PGK isolated from horse, pig, and Bacillus stearothermophilus (rms deviations between equivalent α-carbons in the individual domains < 1.0 Å) but exhibits substantial variations with a previously reported yeast structure (rms deviations between equivalent α-carbons in the individual domains of 2.9−3.2 Å). The most significant tertiary structural differences among the yeast R65Q, equine, porcine, and B. stearothermophilus PGK structures occur in the relative orientations of the two domains. However, the relationships between the observed conformations of PGK are inconsistent with a “hinge-bending” behavior that would close the interdomain cleft. It is proposed that the available structural and biochemical data on PGK may indicate that the basic patch primarily represents the site of anion activation and not the catalytically active binding site for 3-PG.


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http://dx.doi.org/10.1021/bi952500o DOIArticle
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Contact Email Address:phoebe@caltech.edu
Additional Information:© 1996 American Chemical Society. Received October 19, 1995; Revised Manuscript Received January 26, 1996. Supported in part by NIH Grants GM45162 (D.C.R.) and GM37715 (M.T.M.). T.M.M. was supported in part by NIH Predoctoral Training Grant GM07616. The authors thank Dr. Colin F. Blake (Oxford) for providing the unpublished atomic coordinates (porcine PGK, 2.0 Å resolution; equine PGK, 2.5 Å resolution) used in the comparative studies. M.T.M. thanks Dr. J. Kraut (UCSD) and members of his laboratory for hospitality and training in protein crystallization. T.M.M. thanks Ruth Ann Bertsch for invaluable discussions and editing of the manuscript.
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Funding AgencyGrant Number
NIHGM45162
NIHGM37715
NIHGM07616
Record Number:CaltechAUTHORS:20150205-140310848
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:20150205-140310848
Official Citation:Structure of the R65Q Mutant of Yeast 3-Phosphoglycerate Kinase Complexed with Mg-AMP-PNP and 3-Phospho-d-glycerate, Timothy M. McPhillips, Barbara T. Hsu, Mark A. Sherman, Maria T. Mas, and Douglas C. Rees Biochemistry 1996 35 (13), 4118-4127
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ID Code:54435
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Deposited On:08 Feb 2015 03:16
Last Modified:08 Feb 2015 03:16

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