Changes in the proteome of Candida albicans in response to azole, polyene, and echinocandin antifungal agents.

The yeast Candida albicans is an opportunistic human fungal pathogen and the cause of superficial and systemic infections in immunocompromised patients. The classes of antifungal agents most commonly used to treat Candida infections are the azoles, polyenes, and echinocandins. In the present study, we identified changes in C.

Upc2p-associated differential protein expression in Candida albicans.

The gain-of-function mutation G648D in UPC2 causes ERG11 up-regulation and increased fluconazole resistance in Candida albicans. In this study, we performed 2-DE and PMF to identify proteomic alterations in an ERG11-overexpressing fluconazole-resistant C. albicans clinical isolate compared with its fluconazole-susceptible parent strain.

Proteomic analysis of Mrr1p- and Tac1p-associated differential protein expression in azole-resistant clinical isolates of Candida albicans.

Azole resistance in Candida albicans is frequently caused by the overexpression of multi-drug efflux pump genes MDR1, CDR1, and CDR2 due to gain-of-function mutations in the zinc cluster transcription factors Mrr1p and Tac1p. In this study, we performed a comparative proteomic analysis to identify proteins whose expression level is influenced by these transcription factors. Both 2-DE and PMF were used to examine the expression profiles of six pairs of matched C.

Enterotoxigenic Escherichia coli EtpA mediates adhesion between flagella and host cells.

Adhesion to epithelial cells and flagella-mediated motility are critical virulence traits for many Gram-negative pathogens, including enterotoxigenic Escherichia coli (ETEC), a major cause of diarrhoea in travellers and children in developing countries. Many flagellated pathogens export putative adhesins belonging to the two-partner secretion (TPS) family. However, the actual function of these adhesins remains largely undefined.

Proteome cataloging and relative quantification of Francisella tularensis tularensis strain Schu4 in 2D PAGE using preparative isoelectric focusing.

The protein complement of whole cell extract of the bacterium Francisella tularensis tularensis was analyzed using two-dimensional electrophoresis with preparative isoelectric focusing in the first dimension. The format allows the quantification of relative protein abundance by linear densitometry and extends the potential dynamic range of protein detection by as much as an order of magnitude. The relative abundance and rank order of 136 unique proteins identified in F.

High-throughput proteomics processing of proteins in polyacrylamide in a multiwell format.

Processing multiple protein samples from polyacrylamide at significant sensitivity represents a major chokepoint for raising the success rate in high-volume protein identification projects. A multiwell filterplate method for processing proteins in polyacrylamide was optimized for sensitivity using a protein standard. The results demonstrate this process to be a reliable and reproducible method over a range of gel loadings and suitable for the identification of proteins near the threshold of silver stain.

Gene expression profiling of the response of Streptococcus pneumoniae to penicillin.

Objectives: The aim of this study was to identify changes in the gene expression profile of Streptococcus pneumoniae in response to a subinhibitory concentration of penicillin in an effort to better understand mechanisms by which this organism copes with this stress.

Methods: S. pneumoniae serotype 2 strain D39 was grown for 1 h in the presence or absence of penicillin at a concentration equivalent to half the MIC (0.

Proteomic analysis of experimentally induced azole resistance in Candida glabrata.

Objectives: The aim of the present study was to identify changes in the proteome of a laboratory-derived azole-resistant strain of Candida glabrata compared with its susceptible parent strain in an effort to identify proteins that are differentially expressed in association with azole resistance.

Methods: Soluble and membrane protein fractions were isolated from mutant strain F15 (fluconazole MIC>128 mg/L) and parent strain 66032 (fluconazole MIC=16 mg/L) grown to mid-log phase. Soluble proteins were resolved by both two-dimensional (2D) and one-dimensional (1D) polyacrylamide gel electrophoresis (GE) whereas membrane proteins were resolved by 1D GE.

Heterogeneous nuclear ribonucleoprotein P2 is an autoantibody target in mice deficient for Mer, Axl, and Tyro3 receptor tyrosine kinases.

Deficiencies in clearance of apoptotic cells predispose to the development of autoimmune disease. This is evident in mice lacking the receptor tyrosine kinases Tyro3, Axl, and Mer. Deficient mice exhibit an increased abundance of apoptotic cells in tissues and manifest diverse autoimmune conditions.

A mass spectrometric determination of the conformation of dimeric apolipoprotein A-I in discoidal high density lipoproteins.

Discoidal forms of high density lipoproteins (HDL) are critical intermediates between lipid-poor apolipoprotein A-I (apo A-I), the major protein constituent of HDL, and the mature spherical forms that comprise the bulk of circulating particles. Thus, many studies have focused on understanding apoA-I structure in discs reconstituted in vitro. Recent theoretical and experimental work supports a "belt" model for apoA-I in which repeating amphipathic helical domains run parallel to the plane of the lipid disc.

Application of proteomic analysis to the study of azole antifungal resistance in Candida albicans.

The sequencing of the Candida albicans genome and recent refinements in protein resolution and identification techniques have greatly enhanced the application of proteomics for the study of this fungal pathogen. Proteome analysis includes the separation and isolation of proteins by two-dimensional polyacrylamide gel electrophoresis and subsequent protein identification by peptide mass fingerprinting using matrix-assisted laser desorption ionization time-of-flight mass spectrometry. This technique has been used for the study of the proteomes of Candida species in the context of virulence, drug response, and resistance.

A three-dimensional molecular model of lipid-free apolipoprotein A-I determined by cross-linking/mass spectrometry and sequence threading.

Apolipoprotein (apo) A-I, a 243-residue, 28.1-kDa protein is a major mediator of the reverse cholesterol transport (RCT) pathway, a process that may reduce the risk of cardiovascular disease in humans. In plasma, a small fraction of lipid-free or lipid-poor apoA-I is likely a key player in the first step of RCT.

Proteomic analysis of azole resistance in Candida albicans clinical isolates.

Changes in protein expression within a matched set of Candida albicans isolates representing the acquisition of azole resistance were examined by two-dimensional polyacrylamide gel electrophoresis and peptide mass fingerprinting. Proteins differentially expressed in association with azole resistance included Grp2p, Ifd1p, Ifd4p, Ifd5p, and Erg10p, a protein involved in the ergosterol biosynthesis pathway.

Vascular oxygen sensing: detection of novel candidates by proteomics and organ culture.

We have shown that the specific inhibition of hypoxia-induced relaxation by organ culture in porcine coronary arteries can be mimicked by treatment of control vessels with the protein synthesis inhibitor, cycloheximide. We hypothesize that organ culture of vascular smooth muscle results in the decreased expression of proteins that are critical for vascular oxygen sensing. Using two-dimensional gel electrophoresis and mass spectroscopy, we identified such candidate proteins.

The spatial organization of apolipoprotein A-I on the edge of discoidal high density lipoprotein particles: a mass specrometry study.

The three-dimensional structure of human apoA-I on nascent, discoidal HDL particles has been debated extensively over the past 25 years. Recent evidence has demonstrated that the alpha-helical domains of apoA-I are arranged in a belt-like orientation with the long axis of the helices perpendicular to the phospholipid acyl chains on the disc edge. However, experimental information on the spatial relationships between apoA-I molecules on the disc is lacking.

Functional importance of the carboxyl-terminal region of striated muscle tropomyosin.

Striated muscle tropomyosin (TM) interacts with actin and the troponin complex to regulate calcium-mediated muscle contraction. Previous work by our laboratory established that alpha- and beta-TM isoforms elicit physiological differences in sarcomeric performance. Heart myofilaments containing beta-TM exhibit an increased sensitivity to calcium that is associated with a decrease in the rate of relaxation and a prolonged time of relaxation.

Cobalt inhibits the interaction between hypoxia-inducible factor-alpha and von Hippel-Lindau protein by direct binding to hypoxia-inducible factor-alpha.

The hypoxia-inducible factor (HIF) activates the expression of genes that contain a hypoxia response element. The alpha-subunits of the HIF transcription factors are degraded by proteasomal pathways during normoxia but are stabilized under hypoxic conditions. The von Hippel-Lindau protein (pVHL) mediates the ubiquitination and rapid degradation of HIF-alpha (including HIF-1alpha and HIF-2alpha).

Pseudomonas aeruginosa anaerobic respiration in biofilms: relationships to cystic fibrosis pathogenesis.

Recent data indicate that cystic fibrosis (CF) airway mucus is anaerobic. This suggests that Pseudomonas aeruginosa infection in CF reflects biofilm formation and persistence in an anaerobic environment. P.