PDBsum entry 2vyv

Oxidoreductase

PDB id: 2vyv

Contents

Protein chains

330 a.a. *

334 a.a. *

Ligands

FMT ×14

NAD ×4

1GP

Waters ×840

* Residue conservation analysis

PDB id:

2vyv

Name:

Oxidoreductase

Title:

Structure of e.Coli gapdh rat sperm gapdh heterotetramer

Structure:

Glyceraldehyde-3-phosphate dehydrogenase. Chain: a, b, c. Glyceraldehyde-3-phosphate dehydrogenase. Chain: d. Fragment: residues 102-432. Other_details: poly proline n-terminus replaced

Source:

Escherichia coli bl21(de3). Organism_taxid: 469008. Variant: plyss. Rattus norvegicus. Norway rat. Organism_taxid: 10116. Tissue: testis. Other_details: DNA was extracted directly from rat testis

Resolution:

2.38Å R-factor: 0.184 R-free: 0.251

Authors:

J.Frayne,A.Taylor,L.Hall,A.Hadfield

Key ref:

J.Frayne et al. (2009). Structure of insoluble rat sperm glyceraldehyde-3-phosphate dehydrogenase (GAPDH) via heterotetramer formation with Escherichia coli GAPDH reveals target for contraceptive design. J Biol Chem, 284, 22703-22712. PubMed id: 19542219 DOI: 10.1074/jbc.M109.004648

Date:

29-Jul-08 Release date: 05-Aug-08

Protein chains

P0A9B2  (G3P1_ECOLI) -  Glyceraldehyde-3-phosphate dehydrogenase A from Escherichia coli (strain K12)

Seq: 331 a.a.

Struc: 330 a.a.*

Protein chain

Q9ESV6  (G3PT_RAT) -  Glyceraldehyde-3-phosphate dehydrogenase, testis-specific from Rattus norvegicus

Seq: 432 a.a.

Struc: 334 a.a.*

* PDB and UniProt seqs differ at 7 residue positions (black crosses)

Enzyme reactions

• Enzyme class: Chains A, B, C, D: E.C.1.2.1.12 - glyceraldehyde-3-phosphate dehydrogenase (phosphorylating).
• Pathway: Glyceraldehyde-3-phosphate Dehydrogenase (phosphorylating)
• Reaction: D-glyceraldehyde 3-phosphate + phosphate + NAD⁺ = (2R)-3-phospho- glyceroyl phosphate + NADH + H⁺

D-glyceraldehyde 3-phosphate (Bound ligand (Het Group name = 1GP) corresponds exactly) + phosphate (Bound ligand (Het Group name = NAD) corresponds exactly) + NAD(+) = (2R)-3-phospho- glyceroyl phosphate + NADH + H(+)

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1074/jbc.M109.004648

J Biol Chem 284:22703-22712 (2009)

PubMed id: 19542219

Structure of insoluble rat sperm glyceraldehyde-3-phosphate dehydrogenase (GAPDH) via heterotetramer formation with Escherichia coli GAPDH reveals target for contraceptive design.

J.Frayne, A.Taylor, G.Cameron, A.T.Hadfield.

ABSTRACT

Sperm glyceraldehyde-3-phosphate dehydrogenase has been shown to be a successful target for a non-hormonal contraceptive approach, but the agents tested to date have had unacceptable side effects. Obtaining the structure of the sperm-specific isoform to allow rational inhibitor design has therefore been a goal for a number of years but has proved intractable because of the insoluble nature of both native and recombinant protein. We have obtained soluble recombinant sperm glyceraldehyde-3-phosphate dehydrogenase as a heterotetramer with the Escherichia coli glyceraldehyde-3-phosphate dehydrogenase in a ratio of 1:3 and have solved the structure of the heterotetramer which we believe represents a novel strategy for structure determination of an insoluble protein. A structure was also obtained where glyceraldehyde 3-phosphate binds in the P(s) pocket in the active site of the sperm enzyme subunit in the presence of NAD. Modeling and comparison of the structures of human somatic and sperm-specific glyceraldehyde-3-phosphate dehydrogenase revealed few differences at the active site and hence rebut the long presumed structural specificity of 3-chlorolactaldehyde for the sperm isoform. The contraceptive activity of alpha-chlorohydrin and its apparent specificity for the sperm isoform in vivo are likely to be due to differences in metabolism to 3-chlorolactaldehyde in spermatozoa and somatic cells. However, further detailed analysis of the sperm glyceraldehyde-3-phosphate dehydrogenase structure revealed sites in the enzyme that do show significant difference compared with published somatic glyceraldehyde-3-phosphate dehydrogenase structures that could be exploited by structure-based drug design to identify leads for novel male contraceptives.

Selected figure(s)

Figure 7.
Sphere diagram of His-GAPDS highlighting sequence differences between His-GAPDS and human placental GAPDH. Chain D of the His-GAPDS-E. coli GAPDH tetramer is shown in stereo with NAD^+ represented as a blue ball and stick structure. Residue positions that differ in sequence between human GAPDH and GAPDS (blue) are shown as spheres. Residues that differ in sequence in the selectivity cleft are shown as red spheres, and Tyr^98 and Leu^99, which interact with the NAD^+ adenine, are shown as yellow spheres.

Figure 8.
Variability in the P[i] binding pocket.a, transparent surface representation of the human sperm model in the vicinity of the active site, with the loop containing residues 189–192 shown in sticks representation for the human sperm model with carbons in cyan, along with the equivalent loop from human placental GAPDH (1U8F) in light magenta, human liver GAPDH (1ZNQ) in purple, and rabbit muscle GAPDH (1J0X) in white. The Glc-3-P structure is shown with green carbons demonstrating the P[s] site, and a model of 3-chlorolactaldehyde (CL) is shown in the P[i] site, with carbons in blue. b, intersubunit selectivity cleft of the His-GAPDS tetramer is shown in schematic diagram (D, blue; A, yellow; B, red), with the residues that differ in sequence shown as sticks. The equivalent loops in human GAPDH are shown in cyan.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2009, 284, 22703-22712) copyright 2009.

Figures were selected by an automated process.