title: Organophosphate hydrolase in Brevundimonas diminuta is targeted to the periplasmic face of the inner membrane by the twin arginine translocation pathway.
abstract: A twin arginine translocation (Tat) motif, involved in transport of folded proteins across the inner membrane, was identified in the signal peptide of the membrane-associated organophosphate hydrolase (OPH) of Brevundimonas diminuta. Expression of the precursor form of OPH carrying a C-terminal His tag in an opd-negative background and subsequent immunoblotting with anti-His antibodies showed that only the mature form of OPH associated with the membrane and that the precursor form of OPH was entirely found in the cytoplasm. When OPH was expressed without the signal peptide, most of it remained in the cytoplasm, where it was apparently correctly folded and showed activity comparable to that of the membrane-associated OPH encoded by the wild-type opd gene. Amino acid substitutions in the invariant arginine residues of the Tat signal peptide affected both the processing and localization of OPH, confirming a critical role for the Tat system in membrane targeting of OPH in B. diminuta. The localization of OPH to the periplasmic face of the inner membrane in B. diminuta was demonstrated by proteinase K treatment of spheroplasts and also by fluorescence-activated cell sorting analysis of cells expressing OPH-green fluorescent protein fusions with and without an SsrA tag that targets cytoplasmic proteins to the ClpXP protease.
authors.affiliation: Dept. of Animal Sciences, School of Life Sciences, University of Hyderabad, Hyderabad 500 046, India.
authors.forename: Purushotham
authors.initials: P
authors.lastname: Gorla
authors.forename: Jay Prakash
authors.initials: JP
authors.lastname: Pandey
authors.forename: Sunil
authors.initials: S
authors.lastname: Parthasarathy
authors.forename: Mike
authors.initials: M
authors.lastname: Merrick
authors.forename: Dayananda
authors.initials: D
authors.lastname: Siddavattam
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D042063:Caulobacteraceae / Q000201:enzymology*; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D007425:Intracellular Membranes / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D019897:Periplasm / Q000378:metabolism*; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D021381:Protein Transport
chemical_list: D001426:Bacterial Proteins; D026901:Membrane Transport Proteins; C045290:phosphorylphosphatase; D010744:Phosphoric Monoester Hydrolases
references.citation: Annu Rev Microbiol. 2006;60:373-95
references.pmid: 16756481
references.citation: J Bacteriol. 2000 Jul;182(14):4068-76
references.pmid: 10869087
references.citation: J Biol Chem. 1992 Jul 5;267(19):13278-83
references.pmid: 1320014
references.citation: J Mol Biol. 1983 Jun 5;166(4):557-80
references.pmid: 6345791
references.citation: Mol Biol Rep. 2009 Mar;36(3):449-54
references.pmid: 18157614
references.citation: Mol Microbiol. 2003 Jun;48(5):1171-82
references.pmid: 12787347
references.citation: Plasmid. 1987 Sep;18(2):173-7
references.pmid: 2829255
references.citation: Trends Microbiol. 2005 Apr;13(4):175-80
references.pmid: 15817387
references.citation: J Mol Biol. 1986 May 5;189(1):113-30
references.pmid: 3537305
references.citation: Mol Microbiol. 2003 Sep;49(5):1377-90
references.pmid: 12940994
references.citation: J Biotechnol. 2005 Sep 10;118(4):379-85
references.pmid: 15993968
references.citation: Biochem Biophys Res Commun. 2006 Dec 22;351(3):675-81
references.pmid: 17078928
references.citation: Appl Environ Microbiol. 2002 Jul;68(7):3371-6
references.pmid: 12089017
references.citation: J Biol Chem. 2000 Apr 21;275(16):11591-6
references.pmid: 10766774
references.citation: J Biol Chem. 2001 Mar 16;276(11):8159-64
references.pmid: 11099493
references.citation: EMBO J. 2001 May 15;20(10):2472-9
references.pmid: 11350936
references.citation: J Bacteriol. 1989 Dec;171(12):6740-6
references.pmid: 2556372
references.citation: Adv Microb Physiol. 2003;47:187-254
references.pmid: 14560665
references.citation: Science. 1996 Feb 16;271(5251):990-3
references.pmid: 8584937
references.citation: J Bacteriol. 1988 May;170(5):2306-11
references.pmid: 2834339
references.citation: J Biol Chem. 1996 Oct 11;271(41):25582-9
references.pmid: 8810332
references.citation: Appl Environ Microbiol. 1988 Oct;54(10):2586-9
references.pmid: 3202637
references.citation: Curr Opin Microbiol. 2005 Apr;8(2):122-6
references.pmid: 15802241
references.citation: Appl Environ Microbiol. 1986 May;51(5):926-30
references.pmid: 3015022
references.citation: Mol Microbiol. 1996 Nov;22(3):393-404
references.pmid: 8939424
references.citation: Mol Microbiol. 2001 Jan;39(1):47-53
references.pmid: 11123687
references.citation: Biochemistry. 1995 Jun 27;34(25):7973-8
references.pmid: 7794910
references.citation: Arch Microbiol. 2001 Dec;177(1):107-12
references.pmid: 11797051
references.citation: Curr Opin Microbiol. 2005 Apr;8(2):174-81
references.pmid: 15802249
references.citation: J Biol Chem. 2002 Aug 16;277(33):29825-31
references.pmid: 12021272
references.citation: EMBO J. 1998 Jul 1;17(13):3640-50
references.pmid: 9649434
references.citation: Biochemistry. 1994 Dec 20;33(50):15001-7
references.pmid: 7999757
references.citation: Cell. 1998 Apr 3;93(1):93-101
references.pmid: 9546395
references.citation: Gene. 1991 Jan 2;97(1):39-47
references.pmid: 1847347
references.citation: Appl Environ Microbiol. 1988 Feb;54(2):288-93
references.pmid: 3355128
references.citation: Int J Syst Bacteriol. 1994 Jul;44(3):499-510
references.pmid: 8068543
references.citation: Appl Environ Microbiol. 1989 Feb;55(2):289-93
references.pmid: 2541658
references.citation: Can J Microbiol. 1973 Jul;19(7):873-5
references.pmid: 4727806
references.citation: J Bacteriol. 1988 Dec;170(12):5939-42
references.pmid: 3142863
references.citation: J Microsc. 2000 Jun;198(Pt 3):246-59
references.pmid: 10849201
references.citation: Nat Struct Biol. 2000 Jun;7(6):449-55
references.pmid: 10881189
references.citation: Appl Environ Microbiol. 2003 May;69(5):2533-9
references.pmid: 12732518
references.citation: Arch Microbiol. 2002 Dec;178(6):548-53
references.pmid: 12420178