PDBsum entry 1e7y

Oxidoreductase

PDB id: 1e7y

Contents

Protein chain

485 a.a. *

Ligands

BG6

NDP

Metals

_CA

Waters ×131

* Residue conservation analysis

References listed in PDB file

Key reference

Title

An examination of the role of asp-177 in the his-Asp catalytic dyad of leuconostoc mesenteroides glucose 6-Phosphate dehydrogenase: x-Ray structure and ph dependence of kinetic parameters of the d177n mutant enzyme.

Authors

M.S.Cosgrove, S.Gover, C.E.Naylor, L.Vandeputte-Rutten, M.J.Adams, H.R.Levy.

Ref.

Biochemistry, 2000, 39, 15002-15011. [DOI no: 10.1021/bi0014608]

PubMed id

11106478

Abstract

The role of Asp-177 in the His-Asp catalytic dyad of glucose 6-phosphate dehydrogenase from Leuconostoc mesenteroides has been investigated by a structural and functional characterization of the D177N mutant enzyme. Its three-dimensional structure has been determined by X-ray cryocrystallography in the presence of NAD(+) and in the presence of glucose 6-phosphate plus NADPH. The structure of a glucose 6-phosphate complex of a mutant (Q365C) with normal enzyme activity has also been determined and substrate binding compared. To understand the effect of Asp-177 on the ionization properties of the catalytic base His-240, the pH dependence of kinetic parameters has been determined for the D177N mutant and compared to that of the wild-type enzyme. The structures give details of glucose 6-phosphate binding and show that replacement of the Asp-177 of the catalytic dyad with asparagine does not affect the overall structure of glucose 6-phosphate dehydrogenase. Additionally, the evidence suggests that the productive tautomer of His-240 in the D177N mutant enzyme is stabilized by a hydrogen bond with Asn-177; hence, the mutation does not affect tautomer stabilization. We conclude, therefore, that the absence of a negatively charged aspartate at 177 accounts for the decrease in catalytic activity at pH 7.8. Structural analysis suggests that the pH dependence of the kinetic parameters of D177N glucose 6-phosphate dehydrogenase results from an ionized water molecule replacing the missing negative charge of the mutated Asp-177 at high pH. Glucose 6-phosphate binding orders and orients His-178 in the D177N-glucose 6-phosphate-NADPH ternary complex and appears to be necessary to form this water-binding site.

Secondary reference #1

Title

On the mechanism of the reaction catalyzed by glucose 6-Phosphate dehydrogenase.

Authors

M.S.Cosgrove, C.Naylor, S.Paludan, M.J.Adams, H.R.Levy.

Ref.

Biochemistry, 1998, 37, 2759-2767. [DOI no: 10.1021/bi972069y]

PubMed id

9485426

Secondary reference #2

Title

The three-Dimensional structure of glucose 6-Phosphate dehydrogenase from leuconostoc mesenteroides refined at 2.0 a resolution.

Authors

P.Rowland, A.K.Basak, S.Gover, H.R.Levy, M.J.Adams.

Ref.

Structure, 1994, 2, 1073-1087. [DOI no: 10.1016/S0969-2126(94)00110-3]

PubMed id

7881907

Figure 5.
Figure 5. The G6PD dimer viewed from a point just off the dimer axis. Subunit A is shown in red and green in the same orientation as in Figure 2a. Subunit B is shown in magenta and blue. Drawn with MOLSCRIPT [50] and RASTER3D (E Merritt, unpublished program). Figure 5. The G6PD dimer viewed from a point just off the dimer axis. Subunit A is shown in red and green in the same orientation as in [3]Figure 2a. Subunit B is shown in magenta and blue. Drawn with MOLSCRIPT [[4]50] and RASTER3D (E Merritt, unpublished program).

Figure 8.
Figure 8. Differences between the phosphate binding sites in the two subunits. (a) In subunit A there are two well ordered phosphates, PO[4] 2000 and PO[4] 2002. (b) In subunit B, there is only one, PO[4] 2001. Conserved residues are labelled by subunit (A/B) and sequence number. Figure 8. Differences between the phosphate binding sites in the two subunits. (a) In subunit A there are two well ordered phosphates, PO[4] 2000 and PO[4] 2002. (b) In subunit B, there is only one, PO[4] 2001. Conserved residues are labelled by subunit (A/B) and sequence number.

The above figures are reproduced from the cited reference with permission from Cell Press