Show simple item record Chung, Dr Hak Suk en_US Raetz, Christian R. H. en_US 2011-06-21T17:22:09Z 2011-06-21T17:22:09Z 2010 en_US
dc.identifier.citation Chung,Hak Suk;Raetz,Christian R. H.. 2010. Interchangeable Domains in the Kdo Transferases of Escherichia coli and Haemophilus influenzae. Biochemistry 49(19): 4126-4137. en_US
dc.identifier.issn 0006-2960 en_US
dc.description.abstract Kdo(2)-lipid A, a conserved substructure of lipopolysaccharide, plays critical roles in Gram-negative bacterial survival and interaction with host organisms. Inhibition of Kdo biosynthesis in Escherichia coli results in cell death and accumulation of the tetra-acylated precursor lipid IVA. E. coil KdtA (EcKdtA) is a bifunctional enzyme that transfers two Kdo units from two CMP-Kdo molecules to lipid IVA. In contrast, Haemophilia influenzae KdtA (HiKdtA) transfers only one Kdo unit. E. coil CMR300, which lacks Kdo transferase because of a deletion in kdtA, can be rescued to grow in broth at 37 degrees C if multiple copies of msbA are provided in trans. MsbA, the inner membrane transporter for nascent lipopolysaccharide, prefers hexa-acylated to tetra-acylated lipid A, but with the excess MsbA present in CMR300, lipid IVA is efficiently exported to the outer membrane. CMR300 is hypersensitive to hydrophobic antibiotics and bile salts and does not grow at 42 degrees C. Expressing HiKdtA in CMR300 results in the accumulation of Kdo-lipid IVA in place of lipid IVA without suppression of its growth phenotypes at 30 degrees C. EcKdtA restores intact lipopolysaccharide, together with normal antibiotic resistance, detergent resistance, and growth at 42 degrees C. To determine which residues are important for the mono- or bifunctional character of KdtA, protein chimeras were constructed using EcKdtA and HiKdtA. These chimeras, which are catalytically active, were characterized by in vitro assays and in vivo complementation. The N-terminal half of KdtA, especially the first 30 amino acid residues, specifies whether one or two Kdo units are transferred to lipid IVA. en_US
dc.language.iso en_US en_US
dc.publisher AMER CHEMICAL SOC en_US
dc.relation.isversionof doi:10.1021/bi100343e en_US
dc.subject 3-deoxy-d-manno-octulosonic acid transferase en_US
dc.subject bacterial outer-membrane en_US
dc.subject lipid-a en_US
dc.subject lipopolysaccharide en_US
dc.subject biosynthesis en_US
dc.subject gene en_US
dc.subject purification en_US
dc.subject expression en_US
dc.subject phosphorylation en_US
dc.subject cloning en_US
dc.subject biochemistry & molecular biology en_US
dc.title Interchangeable Domains in the Kdo Transferases of Escherichia coli and Haemophilus influenzae en_US
dc.title.alternative en_US
dc.description.version Version of Record en_US 2010-5-18 en_US
duke.description.endpage 4137 en_US
duke.description.issue 19 en_US
duke.description.startpage 4126 en_US
duke.description.volume 49 en_US
dc.relation.journal Biochemistry en_US

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