3-Deoxy-D-manno-octulosonic acid (Kdo) hydrolase identified in Francisella tularensis, Helicobacter pylori, and Legionella pneumophila.
3-Deoxy-D-manno-octulosonic acid (Kdo) is an eight-carbon sugar ubiquitous in Gram-negative bacterial lipopolysaccharides (LPS). Although its biosynthesis is well described, no protein has yet been identified as a Kdo hydrolase. However, Kdo hydrolase enzymatic activity has been detected in membranes of Helicobacter pylori and Francisella tularensis and may be responsible for the removal of side-chain Kdo from the LPS core saccharides. We now report the identification of genes encoding a Kdo hydrolase in F. tularensis Schu S4 and live vaccine strain strains, in H. pylori 26695 strain and in Legionella pneumophila Philadelphia 1 strain. We have renamed the genes kdhA for keto-deoxyoctulosonate hydrolase A. Deletion of kdhA abolished Kdo hydrolase activity in membranes of F. tularensis live vaccine strain. The F. tularensis kdhA mutant synthesized a core oligosaccharide containing a Kdo disaccharide with one of the Kdo residues being a terminal side chain. This side-chain Kdo monosaccharide was absent in the wild-type core oligosaccharide. Expression in Escherichia coli of recombinant KdhA from F. tularensis, H. pylori, and L. pneumophila resulted in a reduction of membrane-associated side-chain Kdo. The identification of this previously faceless enzyme will accelerate study of the biosynthetic basis and biologic impact for postbiosynthetic LPS structural modification.
Chalabaev
Sabina
S
Department of Microbiology and Molecular Genetics, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.
Kim
Tae-Hyun
TH
Ross
Robin
R
Derian
Alec
A
Kasper
Dennis L
DL
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001428:Bacterial Vaccines / Q000235:genetics / Q000378:metabolism; D002462:Cell Membrane / Q000201:enzymology / Q000235:genetics; D005604:Francisella tularensis / Q000201:enzymology* / Q000235:genetics; D017353:Gene Deletion; D006026:Glycoside Hydrolases / Q000235:genetics / Q000378:metabolism*; D016480:Helicobacter pylori / Q000201:enzymology* / Q000235:genetics; D016952:Legionella pneumophila / Q000201:enzymology* / Q000235:genetics; D008070:Lipopolysaccharides / Q000235:genetics / Q000378:metabolism*; D009844:Oligosaccharides / Q000235:genetics / Q000378:metabolism; D013400:Sugar Acids / Q000378:metabolism*; D014613:Vaccines, Attenuated / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D001428:Bacterial Vaccines; D008070:Lipopolysaccharides; D009844:Oligosaccharides; D013400:Sugar Acids; D014613:Vaccines, Attenuated; C002532:2-keto-3-deoxyoctonate; D006026:Glycoside Hydrolases; C053605:KDO alpha-octulopyranoside hydrolase
FEMS Microbiol Lett. 2003 Dec 12;229(2):195-202
14680699
Anal Biochem. 1997 Jan 1;244(1):28-32
9025903
Carbohydr Res. 1990 Mar 25;197:197-204
2189554
Infect Immun. 2007 May;75(5):2591-602
17296751
Nucleic Acids Res. 2004 Jul 1;32(Web Server issue):W327-31
15215404
Biochemistry. 2006 Dec 5;45(48):14427-40
17128982
FEMS Microbiol Immunol. 1992 Oct;5(4):201-10
1419118
J Biol Chem. 1997 Oct 17;272(42):26419-24
9334217
Trends Mol Med. 2008 Aug;14(8):351-60
18606570
Eur J Biochem. 1985 Apr 1;148(1):1-5
2579812
Gene. 1998 May 28;212(1):77-86
9661666
J Biol Chem. 2004 Nov 19;279(47):49470-8
15339914
Curr Drug Discov Technol. 2009 Mar;6(1):19-33
19275539
Am J Bot. 2004 Oct;91(10):1437-45
21652302
J Biol Chem. 2010 Sep 3;285(36):27839-49
20605777
Mol Microbiol. 2010 Nov;78(4):837-52
20659292
Carbohydr Res. 1997 Oct 28;304(1):91-5
9403999
Curr Protoc Protein Sci. 2001 May;Chapter 5:Unit5.2
18429183
J Immunol Methods. 1990 Jan 24;126(1):109-17
2106001
Infect Immun. 1995 Apr;63(4):1183-7
7890370
J Endotoxin Res. 2001;7(1):3-23
11521077
Electrophoresis. 2007 Jul;28(14):2351-7
17578840
Methods Mol Biol. 1995;47:149-54
7550729
Biochemistry. 1978 Feb 21;17(4):645-51
414779
Anal Biochem. 1997 Feb 1;245(1):97-101
9025974
J Biol Chem. 1959 Aug;234(8):1971-5
13672998
J Bacteriol. 2005 May;187(10):3374-83
15866922
J Lipid Res. 2009 Apr;50 Suppl:S103-8
18974037
J Exp Bot. 2008;59(10):2639-47
18503041
Glycoconj J. 1989;6(3):349-53
2562507
Chem Rev. 2002 Feb;102(2):439-69
11841250
J Cell Sci. 2003 Jan 15;116(Pt 2):293-302
12482915
Annu Rev Biochem. 2002;71:635-700
12045108
Eur J Biochem. 2002 Dec;269(24):6112-8
12473106
Infect Immun. 2004 Sep;72(9):5340-8
15322031
Prog Clin Biol Res. 1995;392:113-39
8524918
Planta. 2006 Jun;224(1):222-7
16395581
Plant Cell Physiol. 2007 Mar;48(3):405-13
17234672
J Biol Chem. 1966 Jul 10;241(13):3222-7
5912115
Eur J Biochem. 1997 Dec 1;250(2):608-16
9428716
FEMS Microbiol Lett. 2003 May 28;222(2):273-80
12770718
Mol Microbiol. 2010 Nov;78(4):820-36
20662782
Eur J Biochem. 1989 Jun 1;182(1):153-60
2731548