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PDBsum entry 1v0w
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Contents |
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* Residue conservation analysis
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References listed in PDB file
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Key reference
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Title
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The reaction mechanism of phospholipase d from streptomyces sp. Strain pmf. Snapshots along the reaction pathway reveal a pentacoordinate reaction intermediate and an unexpected final product.
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Authors
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I.Leiros,
S.Mcsweeney,
E.Hough.
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Ref.
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J Mol Biol, 2004,
339,
805-820.
[DOI no: ]
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PubMed id
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Abstract
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Almost all enzyme-catalysed phosphohydrolytic or phosphoryl transfer reactions
proceed through a five-coordinated phosphorus transition state. This is also
true for the phospholipase D superfamily of enzymes, where the active site
usually is made up of two identical sequence repeats of an HKD motif, positioned
around an approximate 2-fold axis, where the histidine and lysine residues are
essential for catalysis. An almost complete reaction pathway has been elucidated
by a series of experiments where crystals of phospholipase D from Streptomyces
sp. strain PMF (PLD(PMF)) were soaked for different times with (i) a soluble
poor, short-chained phospholipid substrate and (ii) with a product. The various
crystal structures were determined to a resolution of 1.35-1.75 A for the
different time-steps. Both substrate and product-structures were determined in
order to identify the different reaction states and to examine if the reaction
actually terminated on formation of phosphatidic acid (the true product of
phospholipase D action) or could proceed even further. The results presented
support the theory that the phospholipase D superfamily shares a common reaction
mechanism, although different family members have very different substrate
preferences and perform different catalytic reactions. Results also show that
the reaction proceeds via a phosphohistidine intermediate and provide
unambiguous identification of a catalytic water molecule, ideally positioned for
apical attack on the phosphorus and consistent with an associative in-line
phosphoryl transfer reaction. In one of the experiments an apparent
five-coordinate phosphorus transition state is observed.
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Figure 1.
Figure 1. The active site of PLD[PMF] unliganded or
phosphate-inhibited. (a) The native state (structure 1). Two
water molecules (OW1 and OW2) occupy the binding site for the
phosphate moiety. (b) Phosphate-inhibited PLD[PMF] (structure
2). Both electron density maps are s[A]-weighted 2mF[o] -DF[c]
maps contoured at 1.5s.
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Figure 8.
Figure 8. The reaction mechanism for PLD[PMF] on a
phosphatidylcholine (PC) substrate. R, Diacylglycerol (DAG); R',
choline. The reaction that takes place when the product
re-enters the active site and the dead-end phosphohistidine is
formed is illustrated below the horizontal line.
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The above figures are
reprinted
by permission from Elsevier:
J Mol Biol
(2004,
339,
805-820)
copyright 2004.
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Secondary reference #1
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Title
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The first crystal structure of a phospholipase d.
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Authors
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I.Leiros,
F.Secundo,
C.Zambonelli,
S.Servi,
E.Hough.
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Ref.
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Structure, 2000,
8,
655-667.
[DOI no: ]
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PubMed id
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Figure 2.
Figure 2. Topology and tertiary structure of PLD. (a)
Overall topology of PLD from Streptomyces sp. strain PMF.
Modified figure from TOPS [46]. The N-terminal 260 residues are
shown in light blue and the remainder of the protein is colored
in dark blue, in order to separate the two domains. (b)
Stereographic overview of the tertiary arrangement of the
protein. Helices are shown in red and strands in yellow. The
protein is viewed from the outer membrane, and the
active-site-bound phosphate can be seen in the center of the
protein. This Figure was created using BOBSCRIPT. (c)
Stereographic presentation of the Ca trace of PLD, made using
BOBSCRIPT. The orientation of the protein is as in (b).
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The above figure is
reproduced from the cited reference
with permission from Cell Press
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