 |
PDBsum entry 2v0h
|
|
|
|
References listed in PDB file
|
 |
|
Key reference
|
|
|
Title
|
|
Characterization of substrate binding and catalysis in the potential antibacterial target n-Acetylglucosamine-1-Phosphate uridyltransferase (glmu).
|
|
|
Authors
|
|
I.Mochalkin,
S.Lightle,
Y.Zhu,
J.F.Ohren,
C.Spessard,
N.Y.Chirgadze,
C.Banotai,
M.Melnick,
L.Mcdowell.
|
|
|
Ref.
|
|
Protein Sci, 2007,
16,
2657-2666.
[DOI no: ]
|
|
|
PubMed id
|
|
|
|
|
|
Abstract
|
|
|
N-Acetylglucosamine-1-phosphate uridyltransferase (GlmU) catalyzes the first
step in peptidoglycan biosynthesis in both Gram-positive and Gram-negative
bacteria. The products of the GlmU reaction are essential for bacterial
survival, making this enzyme an attractive target for antibiotic drug discovery.
A series of Haemophilus influenzae GlmU (hiGlmU) structures were determined by
X-ray crystallography in order to provide structural and functional insights
into GlmU activity and inhibition. The information derived from these structures
was combined with biochemical characterization of the K25A, Q76A, D105A, Y103A,
V223A, and E224A hiGlmU mutants in order to map these active-site residues to
catalytic activity of the enzyme and refine the mechanistic model of the GlmU
uridyltransferase reaction. These studies suggest that GlmU activity follows a
sequential substrate-binding order that begins with UTP binding noncovalently to
the GlmU enzyme. The uridyltransferase active site then remains in an open
apo-like conformation until N-acetylglucosamine-1-phosphate (GlcNAc-1-P) binds
and induces a conformational change at the GlcNAc-binding subsite. Following the
binding of GlcNAc-1-P to the UTP-charged uridyltransferase active site, the
non-esterified oxygen of GlcNAc-1-P performs a nucleophilic attack on the
alpha-phosphate group of UTP. The new data strongly suggest that the mechanism
of phosphotransfer in the uridyltransferase reaction in GlmU is primarily
through an associative mechanism with a pentavalent phosphate intermediate and
an inversion of stereochemistry. Finally, the structural and biochemical
characterization of the uridyltransferase active site and catalytic mechanism
described herein provides a basis for the structure-guided design of novel
antibacterial agents targeting GlmU activity.
|
|
|
|
|
|
|
|
Figure 2.
Figure 2. hiGlmU uridyltransferase active site. (A) Stereoview of the (Fo-Fc) OMIT electron density maps of UDP-GlcNAc, UDP, and uridine bound to the
|
|
Figure 4.
Figure 4. Structural insights into the mechanism of uridylation. (A) View of the GlmU uridyltransferase active site (open conformation) in the UDP-bound
|
|
|
|
|
|
The above figures are
reprinted
by permission from the Protein Society:
Protein Sci
(2007,
16,
2657-2666)
copyright 2007.
|
|
|
|
|
|
|
