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PDBsum entry 2iu8
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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 and reactivity of lpxd, The n-Acyltransferase of lipid a biosynthesis.
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Authors
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L.Buetow,
T.K.Smith,
A.Dawson,
S.Fyffe,
W.N.Hunter.
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Ref.
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Proc Natl Acad Sci U S A, 2007,
104,
4321-4326.
[DOI no: ]
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PubMed id
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Abstract
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The external layer of the Gram-negative bacterial outer membrane is primarily
composed of a protective, selectively permeable LPS. The biosynthesis of LPS
relies on UDP-3-O-acyl-glucosamine N-acyltransferase (LpxD), which transfers
3-hydroxy-arachidic acid from acyl carrier protein to the 2' amine of
UDP-3-O-myristoyl glucosamine in Chlamydia trachomatis. Our crystallographic
study reveals that LpxD is a homotrimer, each subunit of which is constructed
from a novel combination of an N-terminal uridine-binding domain, a core
lipid-binding domain, and a C-terminal helical extension. Highly conserved
residues dominate nucleotide binding. Phe-43 and Tyr-49 form pi-stacking
interactions with uracil, and Asn-46 and His-284 form hydrogen bonds with the
phosphate groups. These interactions place the glucosamine moiety at the
catalytic center formed by two adjacent subunits. Here His-247 and His-284
contribute to a mechanism involving nucleophilic attack by the amine of one
substrate on the carbonyl carbon of an acyl carrier protein thioester conjugate.
Serendipitously, our study reveals a fatty acid (FA) binding groove near the
catalytic center. MS elucidated the presence of a FA mixture binding to LpxD,
with palmitic acid the most prevalent. The placement of UDP-N-acetylglucosamine
and the FA provides details of N-acyltransferase ligand interactions and allows
for a description of structure and reactivity at an early stage of LPS assembly.
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Figure 2.
Fig. 2. Structure of LpxD. (A) Ribbon diagram of a subunit.
The UBD is yellow, the LBD is blue, loops are magenta, and the
HE is red. Selected elements of secondary structure are labeled,
and the coils are numbered. (B) The trimer. The view is parallel
to the noncrystallographic symmetry threefold axis. Subunits are
colored gray, wheat, and slate, and the domains of the gray
subunit are labeled. UDP-GlcNAc (complex II) is represented by
spheres, and palmitic acid is represented by sticks. The atoms
of the ligands are colored as follows: C, green; N, blue; O,
red; P, yellow. (C) Orthogonal view of the trimer. (D) Primary
and secondary structure.  -strands are depicted by
arrows, and  -helices are depicted
by cylinders. Colors are as described in A. Disordered residues
at the C terminus are marked by dots. Highly conserved residues
(>60% identity in 85 sequences) are highlighted in gray, and
strictly conserved residues (100% identity) are in black. Green
stars and circles indicate residues that interact with
UDP-GlcNAc and palmitic acid, respectively. Salmon boxes
represent sites of conditionally lethal point mutations in E.
coli and S. typhimurium LpxD.
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Figure 4.
Fig. 4. LpxD–FA complex. (A) Identification of bound FA.
GC-MS analysis, chain length, and saturation states are
indicated, and the key shows the relative percentages. (B)
Surface view of conserved residues with ligands depicted as
sticks. Palmitic acid is colored cyan, and UDP-GlcNAc is colored
according to atom type: C, white; N, blue; O, red. Conserved
residues are colored by type; basic residues are blue with the
exception of His-247 and His-284, which are colored green.
Acidic residues are red, aromatic residues are salmon, glycine
residues are yellow, polar residues (Asn, Gln, Ser, Thr, and
Cys) are slate blue, and aliphatic residues (Ala, Ile, Val, Leu,
Met, and Pro) are magenta. Colored residues that form part of
the FA and UDP-GlcNAc binding pockets include Gly-262, Gly-280,
Gly-265, Ala-246, Ile-263, Ala-264, Asp-240, and Gln-244.
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