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PDBsum entry 2vyv
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Oxidoreductase
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PDB id
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2vyv
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References listed in PDB file
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Key reference
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Title
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Structure of insoluble rat sperm glyceraldehyde-3-Phosphate dehydrogenase (gapdh) via heterotetramer formation with escherichia coli gapdh reveals target for contraceptive design.
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Authors
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J.Frayne,
A.Taylor,
G.Cameron,
A.T.Hadfield.
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Ref.
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J Biol Chem, 2009,
284,
22703-22712.
[DOI no: ]
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PubMed id
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Abstract
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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.
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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.
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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.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2009,
284,
22703-22712)
copyright 2009.
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