1ppx
From Proteopedia
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'''Solution Structure of the MutT Pyrophosphohydrolase Complexed with Mg(2+) and 8-oxo-dGMP, a Tightly-bound Product''' | '''Solution Structure of the MutT Pyrophosphohydrolase Complexed with Mg(2+) and 8-oxo-dGMP, a Tightly-bound Product''' | ||
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[[Category: Mildvan, A S.]] | [[Category: Mildvan, A S.]] | ||
[[Category: Saraswat, V.]] | [[Category: Saraswat, V.]] | ||
| - | [[Category: | + | [[Category: Mutator protein]] |
| - | [[Category: | + | [[Category: Mutt pyrophosphohydrolase-metal-product complex]] |
| - | [[Category: | + | [[Category: Nucleoside triphosphate pyrophosphohydrolase]] |
| - | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sat May 3 05:21:03 2008'' | |
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on | + | |
Revision as of 02:21, 3 May 2008
Solution Structure of the MutT Pyrophosphohydrolase Complexed with Mg(2+) and 8-oxo-dGMP, a Tightly-bound Product
Overview
To learn the structural basis for the unusually tight binding of 8-oxo-nucleotides to the MutT pyrophosphohydrolase of Escherichia coli (129 residues), the solution structure of the MutT-Mg(2+)-8-oxo-dGMP product complex (K(D) = 52 nM) was determined by standard 3-D heteronuclear NMR methods. Using 1746 NOEs (13.5 NOEs/residue) and 186 phi and psi values derived from backbone (15)N, Calpha, Halpha, and Cbeta chemical shifts, 20 converged structures were computed with NOE violations <or=0.25 A and total energies <or=450 kcal/mol. The pairwise root-mean-square deviations (RMSD) of backbone N, Calpha, and C' atoms for the secondary structured regions and for all residues of the 20 structures are 0.65 and 0.98 A, respectively, indicating a well-defined structure. Further refinement using residual dipolar coupling from 53 backbone N-H vectors slightly improved the RMSD values to 0.49 and 0.84 A, respectively. The secondary structures, which consisted of two alpha-helices and a five-stranded mixed beta-sheet, were indistinguishable from those of free MutT and of MutT in the quaternary MutT-Mg(2+)-(H(2)O)-AMPCPP-Mg(2+) complex. Comparisons of these three tertiary structures showed a narrowing of the hydrophobic nucleotide-binding cleft in the 8-oxo-dGMP complex resulting from a 2.5-4.5 A movement of helix I and a 1.5 A movement of helix II and loop 4 toward the cleft. The binding of 8-oxo-dGMP to MutT-Mg(2+) buries 71-78% of the surface area of the nucleotide. The 10(3.7)-fold weaker binding substrate analogue Mg(2+)-AMPCPP induced much smaller changes in tertiary structure, and MutT buried only 57% of the surface of the AMP moiety of AMPCPP. Formation of the MutT-Mg(2+)-8-oxo-dGMP complex slowed the backbone NH exchange rates of 45 residues of the enzyme by factors of 10(1)-10(6) as compared with the MutT-Mg(2+) and the MutT-Mg(2+)-dGMP complexes, suggesting a more compact structure when 8-oxo-dGMP is bound. The 10(4.6)-fold weaker binding of dGMP to MutT-Mg(2+) (K(D) = 1.8 mM) slowed the backbone exchange rates of only 20 residues and by smaller factors of approximately 10. Hence, the high affinity of MutT-Mg(2+) for 8-oxo-dGMP likely results from widespread ligand-induced conformation changes that narrow the nucleotide binding site and lower the overall free energy of the enzyme-product complex. Specific hydrogen bonding of the purine ring of 8-oxo-dGMP by the side chains of Asn-119 and Arg-78 may also contribute.
About this Structure
1PPX is a Single protein structure of sequence from Escherichia coli. Full crystallographic information is available from OCA.
Reference
Solution structure and NH exchange studies of the MutT pyrophosphohydrolase complexed with Mg(2+) and 8-oxo-dGMP, a tightly bound product., Massiah MA, Saraswat V, Azurmendi HF, Mildvan AS, Biochemistry. 2003 Sep 2;42(34):10140-54. PMID:12939141 Page seeded by OCA on Sat May 3 05:21:03 2008
