title: Lipid-modified azurin of Neisseria meningitidis is a copper protein localized on the outer membrane surface and not regulated by FNR.
abstract: The laz gene of Neisseria meningitidis is predicted to encode a lipid-modified azurin (Laz). Laz is very similar to azurin, a periplasmic protein, which belongs to the copper-containing proteins in the cupredoxin superfamily. In other bacteria, azurin is an electron donor to nitrite reductase, an important enzyme in the denitrifying process. It is not known whether Laz could function as an electron transfer protein in this important pathogen. Laz protein was heterologously expressed in Escherichia coli and purified. Electrospray mass spectrometry indicated that the Laz protein contains one copper ion. Laz was shown to be redox-active in the presence of its redox center copper ion. When oxidized, Laz exhibits an intense blue colour and absorbs visible light around 626 nm. The absorption is lost when exposed to diethyldithiocarbamate, a copper chelating agent. Polyclonal antibodies were raised against purified Laz for detecting expression of Laz under different growth conditions and to determine the orientation of Laz on the outer membrane. The expression of Laz under microaerobic and microaerobic denitrifying conditions was slightly higher than that under aerobic conditions. However, the expression of Laz was similar between the wild type strain and an fnr mutant, suggesting that Fumarate/Nitrate reduction regulator (FNR) does not regulate the expression of Laz despite the presence of a partial FNR box upstream of the laz gene. We propose that some Laz protein is exposed on the outer membrane surface of N. meningitidis as the αLaz antibodies can increase killing by complement in a capsule deficient N. meningitidis strain, in a dose-dependent fashion.
authors: Department of Biology (Area 10), University of York, Heslington, York, YO10 5YW, UK, manu.deeudom@cmu.ac.th.
authors: Manu
authors: M
authors: Deeudom
authors: Wilhemina
authors: W
authors: Huston
authors: James W B
authors: JW
authors: Moir
mesh_terms: D000332:Aerobiosis; D000595:Amino Acid Sequence; D001400:Azurin / Q000032:analysis* / Q000235:genetics; D001425:Bacterial Outer Membrane Proteins / Q000032:analysis* / Q000737:chemistry* / Q000235:genetics; D003300:Copper / Q000032:analysis*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D017353:Gene Deletion; D015870:Gene Expression; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D009345:Neisseria meningitidis / Q000737:chemistry* / Q000235:genetics; D010084:Oxidation-Reduction; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D021241:Spectrometry, Mass, Electrospray Ionization; D014157:Transcription Factors / Q000235:genetics
chemical_list: D001425:Bacterial Outer Membrane Proteins; D011994:Recombinant Proteins; D014157:Transcription Factors; D001400:Azurin; D003300:Copper