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J Biol Chem. 2014 Aug 29;289(35):24059-68. doi: 10.1074/jbc.M114.589986. Epub 2014 Jul 14.

Structural basis of lipid binding for the membrane-embedded tetraacyldisaccharide-1-phosphate 4'-kinase LpxK.

Author information

1
From the Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710 and.
2
the Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee 37205.
3
From the Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710 and peizhou@biochem.duke.edu.

Abstract

The membrane-bound tetraacyldisaccharide-1-phosphate 4'-kinase, LpxK, catalyzes the sixth step of the lipid A (Raetz) biosynthetic pathway and is a viable antibiotic target against emerging Gram-negative pathogens. We report the crystal structure of lipid IVA, the LpxK product, bound to the enzyme, providing a rare glimpse into interfacial catalysis and the surface scanning strategy by which many poorly understood lipid modification enzymes operate. Unlike the few previously structurally characterized proteins that bind lipid A or its precursors, LpxK binds almost exclusively to the glucosamine/phosphate moieties of the lipid molecule. Steady-state kinetic analysis of multiple point mutants of the lipid-binding pocket pinpoints critical residues involved in substrate binding, and characterization of N-terminal helix truncation mutants uncovers the role of this substructure as a hydrophobic membrane anchor. These studies make critical contributions to the limited knowledge surrounding membrane-bound enzymes that act upon lipid substrates and provide a structural template for designing small molecule inhibitors targeting this essential kinase.

KEYWORDS:

Antibiotic; Crystal Structure; Enzyme; Lipid A; Lipid Kinase; Lipid-binding Protein; Lipopolysaccharide (LPS); Membrane Protein

PMID:
25023290
PMCID:
PMC4148839
DOI:
10.1074/jbc.M114.589986
[Indexed for MEDLINE]
Free PMC Article

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