Identification of Rv3852 as a nucleoid-associated protein in Mycobacterium tuberculosis.
Tuberculosis remains the major cause of mortality due to a bacterial pathogen, Mycobacterium tuberculosis. The molecular mechanisms of infection and persistence have not been completely elucidated for this pathogen. Studies involving nucleoid-associated proteins (NAPs), which have been related to the control and influence of virulence genes in pathogenic bacteria, can help unveil the virulence process of M. tuberculosis. Here, we describe the initial characterization of an ORF for an M. tuberculosis putative NAP. The Rv3852 gene was cloned and expressed, and its product purified to homogeneity. A qualitative protein-DNA binding assay was carried out by gel-retardation and the protein affinity for specific DNA sequences was assessed quantitatively by surface plasmon resonance (SPR). A stoichiometry of 10 molecules of monomeric protein per molecule of DNA was determined. The monophasic apparent dissociation rate constant values increased to a saturable level as a function of protein concentration, yielding two limiting values for the molecular recognition of proU2 DNA. A protein-DNA binding mechanism is proposed. In addition, functional complementation studies with an Escherichia coli hns mutant reinforce the likelihood that the Rv3852 protein represents a novel NAP in M. tuberculosis.
Werlang
Isabel C R
ICR
Programa de Pós-Graduação em Biologia Celular e Molecular, Universidade Federal do Rio Grande do Sul, Av. Bento Gonçalves 9500, Porto Alegre, RS 91501-970, Brazil.|Centro de Pesquisas em Biologia Molecular e Funcional, Instituto Nacional de Ciência e Tecnologia em Tuberculose, Pontifícia Universidade Católica do Rio Grande do Sul, Av. Ipiranga 6681, Porto Alegre, RS 90619-900, Brazil.
Schneider
Cristopher Z
CZ
Centro de Pesquisas em Biologia Molecular e Funcional, Instituto Nacional de Ciência e Tecnologia em Tuberculose, Pontifícia Universidade Católica do Rio Grande do Sul, Av. Ipiranga 6681, Porto Alegre, RS 90619-900, Brazil.
Mendonça
Jordana D
JD
Centro de Pesquisas em Biologia Molecular e Funcional, Instituto Nacional de Ciência e Tecnologia em Tuberculose, Pontifícia Universidade Católica do Rio Grande do Sul, Av. Ipiranga 6681, Porto Alegre, RS 90619-900, Brazil.
Palma
Mario S
MS
Laboratório de Biologia Estrutural e Zooquímica, Centro de Estudos de Insetos Sociais, Departamento de Biologia, Instituto de Biociências, Universidade Estadual Paulista, Rio Claro, SP 13506-900, Brazil.
Basso
Luiz A
LA
Centro de Pesquisas em Biologia Molecular e Funcional, Instituto Nacional de Ciência e Tecnologia em Tuberculose, Pontifícia Universidade Católica do Rio Grande do Sul, Av. Ipiranga 6681, Porto Alegre, RS 90619-900, Brazil.
Santos
Diógenes S
DS
Centro de Pesquisas em Biologia Molecular e Funcional, Instituto Nacional de Ciência e Tecnologia em Tuberculose, Pontifícia Universidade Católica do Rio Grande do Sul, Av. Ipiranga 6681, Porto Alegre, RS 90619-900, Brazil.
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004587:Electrophoresis, Agar Gel; D024202:Electrophoretic Mobility Shift Assay; D004926:Escherichia coli / Q000378:metabolism; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000378:metabolism; D016366:Open Reading Frames; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D011994:Recombinant Proteins