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Structural features responsible for the biological stability of Histoplasma's virulence factor CBP.

ABSTRACT: The virulence factor CBP is the most abundant protein secreted by Histoplasma capsulatum, a pathogenic fungus that causes histoplasmosis. Although the biochemical function and pathogenic mechanism of CBP are unknown, quantitative Ca (2+) binding measurements indicate that CBP has a strong affinity for calcium ( K D = 6.45 +/- 0.4 nM). However, no change in structure was observed upon binding of calcium, prompting a more thorough investigation of the molecular properties of CBP with respect to self-association, secondary structure, and stability. Over a wide range of pH values and salt concentrations, CBP exists predominantly as a stable, noncovalent homodimer in both its calcium-free and -bound states. Solution-state NMR and circular dichroism (CD) measurements indicated that the protein is largely alpha-helical, and its secondary structure content changes little over the range of pH values encountered physiologically. ESI-MS revealed that the six cysteine residues of CBP are involved in three intramolecular disulfide bonds that help maintain a highly protease resistant structure. Thermally and chemically induced denaturation studies indicated that unfolding of disulfide-intact CBP is reversible and provided quantitative measurements of protein stability. This disulfide-linked, protease resistant, homodimeric alpha-helical structure of CBP is likely to be advantageous for a virulence factor that must survive the harsh environment within the phagolysosomes of host macrophages.

SUBMITTER: Beck MR

PROVIDER: S-EPMC3124767 | biostudies-literature | 2008 Apr

REPOSITORIES: biostudies-literature

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Structural features responsible for the biological stability of Histoplasma's virulence factor CBP.

Beck Moriah R MR DeKoster Gregory T GT Hambly David M DM Gross Michael L ML Cistola David P DP Goldman William E WE

Biochemistry 20080325 15

The virulence factor CBP is the most abundant protein secreted by Histoplasma capsulatum, a pathogenic fungus that causes histoplasmosis. Although the biochemical function and pathogenic mechanism of CBP are unknown, quantitative Ca (2+) binding measurements indicate that CBP has a strong affinity for calcium ( K D = 6.45 +/- 0.4 nM). However, no change in structure was observed upon binding of calcium, prompting a more thorough investigation of the molecular properties of CBP with respect to se ...[more]

PMID: 18361504

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Project description:BackgroundAn important goal in post-genomic research is discovering the network of interactions between transcription factors (TFs) and the genes they regulate. We have previously reported the development of a supervised-learning approach to TF target identification, and used it to predict targets of 104 transcription factors in yeast. We now include a new sequence conservation measure, expand our predictions to include 59 new TFs, introduce a web-server, and implement an improved ranking method to reveal the biological features contributing to regulation. The classifiers combine 8 genomic datasets covering a broad range of measurements including sequence conservation, sequence overrepresentation, gene expression, and DNA structural properties.Principal findings(1) Application of the method yields an amplification of information about yeast regulators. The ratio of total targets to previously known targets is greater than 2 for 11 TFs, with several having larger gains: Ash1(4), Ino2(2.6), Yaf1(2.4), and Yap6(2.4). (2) Many predicted targets for TFs match well with the known biology of their regulators. As a case study we discuss the regulator Swi6, presenting evidence that it may be important in the DNA damage response, and that the previously uncharacterized gene YMR279C plays a role in DNA damage response and perhaps in cell-cycle progression. (3) A procedure based on recursive-feature-elimination is able to uncover from the large initial data sets those features that best distinguish targets for any TF, providing clues relevant to its biology. An analysis of Swi6 suggests a possible role in lipid metabolism, and more specifically in metabolism of ceramide, a bioactive lipid currently being investigated for anti-cancer properties. (4) An analysis of global network properties highlights the transcriptional network hubs; the factors which control the most genes and the genes which are bound by the largest set of regulators. Cell-cycle and growth related regulators dominate the former; genes involved in carbon metabolism and energy generation dominate the latter.ConclusionPostprocessing of regulatory-classifier results can provide high quality predictions, and feature ranking strategies can deliver insight into the regulatory functions of TFs. Predictions are available at an online web-server, including the full transcriptional network, which can be analyzed using VisAnt network analysis suite.

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Structural features responsible for the biological stability of Histoplasma's virulence factor CBP.

Publications

Structural features responsible for the biological stability of Histoplasma's virulence factor CBP.

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