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PubMed2026

H-NOX and NosP Regulate Flagellar Protein and Virulence Factor Production in Vibrio cholerae.

The ability of Vibrio cholerae to transition between motile and sessile forms in the environment and in the host is critical to its survival and virulence. The molecular cues, sensor proteins, and signaling pathways mediating these transitions are highly complex and often overlapping. Nevertheless, a detailed understanding of them is critical for understanding the persistence and pathogenesis of this deadly pathogen. Nitric oxide (NO) functions as an important signaling molecule in many bacteria, affecting biofilm formation, motility, and virulence, often through interaction with heme protein sensors. The genome of V. cholerae encodes two such sensors called H-NOX and NosP. Here we constructed a Δhnox/nosP mutant and employed a multi-omics methodology that combines tandem-mass-tag (TMT)-based quantitative proteomics, phosphoproteomics, and targeted metabolomics to investigate the function of these sensors. A set of 258 proteins was differentially expressed in the mutant that included many proteins involved in flagellar biosynthesis and motility as well as critical virulence factors, iron acquisition systems, and metabolic enzymes. Many of the identified genes are also part of the ferric uptake regulator (Fur) regulon and iron-dependent transcriptional repression of several Fur targets was disrupted. Phosphoproteomics analysis also revealed proteins involved in motility and virulence as differentially phosphorylated in the mutant strain. In most cases, these phosphoproteins have not been previously observed and provide a wealth of new targets for investigating mechanisms of V. cholerae signaling. Taken together, this work illustrates a role for H-NOX and NosP in promoting factors important for infection while suppressing those important for environmental survival, suggesting a function in priming the organism for infection and/or maintaining the infectious phenotype.

Journal Article↗

PubMed2026

Deciphering S-nitrosylation-regulated metabolic networks in postmortem beef based on label-free modificomics: Identification of ferroptosis as a novel quality-related pathway.

This study elucidated the molecular mechanisms of S-nitrosylation on postmortem beef metabolism and quality based on the label-free modificomics. Varying degrees of S-nitrosylation were exogenously induced in beef semimembranosus (SM) muscle. Results indicated that a high S-nitrosylation level significantly increased beef pH and Warner-Bratzler shear force (WBSF) while reducing centrifugal loss (P < 0.05). A total of 828 S-nitrosylated proteins and 1458 modification sites were identified, of which 114 sites on 81 proteins (DSNPs) exhibited differential modification abundance, representing an increase of 125% compared with previous proteomics studies. DSNPs were mainly involved in glycolysis, the tricarboxylic acid cycle, oxidative phosphorylation, calcium signaling, cell structure, and ferroptosis. Notably, this study provides the first evidence in postmortem muscle that S-nitrosylation regulates key ferroptosis-related proteins, including ACSL, CP, and TF, offering new insights into the link between S-nitrosylation and the ferroptosis pathway in meat. Correlation analysis demonstrated that TF was significantly negatively correlated with pH and WBSF, but positively correlated with centrifugal loss (P < 0.05). Collectively, protein S-nitrosylation critically modulates postmortem beef quality through the coordinated regulation of multiple metabolic processes. More importantly, the identification of ferroptosis as a S-nitrosylation-sensitive pathway provides a new perspective for regulating meat quality through protein post-translational modifications.

Animals↗

PubMed2006

Proteomic profiling and identification of immunodominant spore antigens of Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis.

Differentially expressed and immunogenic spore proteins of the Bacillus cereus group of bacteria, which includes Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis, were identified. Comparative proteomic profiling of their spore proteins distinguished the three species from each other as well as the virulent from the avirulent strains. A total of 458 proteins encoded by 232 open reading frames were identified by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry analysis for all the species. A number of highly expressed proteins, including elongation factor Tu (EF-Tu), elongation factor G, 60-kDa chaperonin, enolase, pyruvate dehydrogenase complex, and others exist as charge variants on two-dimensional gels. These charge variants have similar masses but different isoelectric points. The majority of identified proteins have cellular roles associated with energy production, carbohydrate transport and metabolism, amino acid transport and metabolism, posttranslational modifications, and translation. Novel vaccine candidate proteins were identified using B. anthracis polyclonal antisera from humans postinfected with cutaneous anthrax. Fifteen immunoreactive proteins were identified in B. anthracis spores, whereas 7, 14, and 7 immunoreactive proteins were identified for B. cereus and in the virulent and avirulent strains of B. thuringiensis spores, respectively. Some of the immunodominant antigens include charge variants of EF-Tu, glyceraldehyde-3-phosphate dehydrogenase, dihydrolipoamide acetyltransferase, Delta-1-pyrroline-5-carboxylate dehydrogenase, and a dihydrolipoamide dehydrogenase. Alanine racemase and neutral protease were uniquely immunogenic to B. anthracis. Comparative analysis of the spore immunome will be of significance for further nucleic acid- and immuno-based detection systems as well as next-generation vaccine development.

Antigens, Bacterial↗

PubMed2007

A proteomic analysis of Psychrobacter articus 273-4 adaptation to low temperature and salinity using a 2-D liquid mapping approach.

Psychrobacter 273-4 was isolated from a 20,000-40,000-year-old Siberian permafrost core, which is characterized by low temperature, low water activity, and high salinity. To explore how 273-4 survives in the permafrost environment, proteins in four 273-4 samples cultured at 4 and 22 degrees C in media with and without 5% sodium chloride were profiled and comparatively studied using 2-D HPLC and MS. The method used herein involved fractionation via a pH gradient using chromatofocusing followed by nonporous silica (NPS) RP-HPLC and on-line electrospray mass mapping. It was observed that 33 proteins were involved in the adaptation to low temperature in the cells grown in the nonsaline media while there were only 14 proteins involved in the saline media. There were 45 proteins observed differentially expressed in response to salt at 22 degrees C while there were 22 proteins at 4 degrees C. In addition, 5% NaCl and 4 degrees C showed a combination effect on protein expression. A total of 56 proteins involved in the adaptation to low temperature and salt were identified using MS and database searching. The differentially expressed proteins were classified into different functional categories where the response of the regulation system to stress appears to be very elaborate. The evidence shows that the adaptation of 273-4 is based primarily on the control of translation and transcription, the synthesis of proteins (chaperones) to facilitate RNA and protein folding, and the regulation of metabolic pathways.

Acclimatization↗

PubMedDate not supplied

Metabolic labeling of human primary retinal pigment epithelial cells for accurate comparative proteomics.

Metabolic labeling was evaluated, using both 13C6-Arg and 13C6, 15N2-Lys amino acids, for a primary human retinal pigment epithelial cell (hRPE) culture prepared from an autopsy eye of an 81 year old donor. Satisfactory incorporation (>90%) was achieved with both stable isotope labeled amino acids after four passages (roughly 7 population doublings). The degree of incorporation was found to be efficient with both amino acids as well as in different proteins. The presence of 10% whole serum in the culture medium did not interfere with the incorporation of the exogenous stable isotope labeled amino acids. Metabolic labeling of these human primary retinal pigment epithelial cells was further tested to quantify protein ratios between proliferating and resting cells using a combination of 2-DG and MALDI-TOF-TOF/MS analysis. Using computational data processing and analysis, we obtained accurate protein ratio measurement for every single identified protein (156 proteins) in the 2-Dg array. Of these 156 proteins, 12 proteins were found significantly increased in dividing versus resting cells by at least a factor of 1.5 while 13 other proteins were found increased in resting versus dividing cells by at least the same fold. Most of these differentially expressed proteins are directly involved in cell proliferation, protein synthesis, and actin-remodeling and differentiation.

Aged↗

PubMed2006

Proteomic profiling of primary breast cancer predicts axillary lymph node metastasis.

To determine if protein expression in primary breast cancers can predict axillary lymph node (ALN) metastasis, we assessed differences in protein expression between primary breast cancers with and without ALN metastasis using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS). Laser capture microdissection was performed on invasive breast cancer frozen sections from 65 patients undergoing resection with sentinel lymph node (SLN) or level I and II ALN dissection. Isolated proteins from these tumors were applied to immobilized metal affinity capture (IMAC-3) ProteinChip arrays and analyzed by SELDI-TOF-MS to generate unique protein profiles. Correlations between unique protein peaks and histologically confirmed ALN status and other known clinicopathologic factors were examined using ANOVA and multivariate logistic regression. Two metal-binding polypeptides at 4,871 and 8,596 Da were identified as significant risk factors for nodal metastasis (P = 0.034 and 0.015, respectively) in a multivariate analysis. Lymphovascular invasion (LVI) was the only clinicopathologic factor predictive of ALN metastasis (P = 0.0038). In a logistic regression model combining the 4,871 and 8,596 Da peaks with LVI, the area under the receiver operating characteristic curve was 0.87. Compared with patients with negative ALN, those with > or =2 positive ALN or non-SLN metastases were significantly more likely to have an increased peak at 4,871 Da (P = 0.016 and 0.0083, respectively). ProteinChip array analysis identified differential protein peaks in primary breast cancers that predict the presence and number of ALN metastases and non-SLN status.

Biopsy↗

PubMed2006

A novel histology-directed strategy for MALDI-MS tissue profiling that improves throughput and cellular specificity in human breast cancer.

We describe a novel tissue profiling strategy that improves the cellular specificity and analysis throughput of protein profiles obtained by direct MALDI analysis. The new approach integrates the cellular specificity of histology, the accuracy and reproducibility of robotic liquid dispensing, and the speed and objectivity of automated spectra acquisition. Traditional methodologies for preparing and analyzing tissue samples rely heavily on manual procedures, which for various reasons discussed, restrict cellular specificity and sample throughput. Here, a robotic spotter deposits micron-sized droplets of matrix precisely onto foci of normal mammary epithelium, ductal carcinoma in situ, invasive mammary cancer, and peritumoral stroma selected by a pathologist from high resolution histological images of sectioned human breast cancer samples. The location of each matrix spot was then determined and uploaded into the instrument to facilitate automated profile acquisition by MALDI-TOF. In the example shown, the different lesions were clearly differentiated using mass profiling. Further, the workflow permits a visual projection of any information produced from the profile analyses directly on the histological image for a unique combination of proteomic and histological assessment of sample regions. The higher performance characteristics offered by the new workflow promises to be a significant advancement toward the next generation of tissue profiling studies.

Adult↗

PubMed2005

Proteome investigation of the global regulatory role of sigma 54 in response to gentisate induction in Pseudomonas alcaligenes NCIMB 9867.

Pseudomonas alcaligenes NCIMB 9867 (strain P25X) utilizes the gentisate pathway for the degradation of aromatic hydrocarbons. The gene encoding the alternative sigma (sigma) factor sigma(54), rpoN, was cloned from strain P25X and a rpoN knock-out strain, designated G54, was constructed by insertional inactivation with a kanamycin resistance gene cassette. The role of sigma(54) in the physiological response of P. alcaligenes P25X to gentisate induction was assessed by comparing the global protein expression profiles of the wild-type P25X with the rpoN mutant strain G54. Analysis of two-dimensional polyacrylamide gel electrophoresis gels showed that 39 out of 355 prominent protein spots exhibited differential expression as a result of the insertional inactivation of rpoN. Identification of the protein spots by matrix-assisted laser desorption/ionization-time of flight/time of flight revealed a wide diversity of proteins that are affected by the sigma(54) mutation, the largest group being proteins that are involved in carbon metabolism. The strictly inducible gentisate 1,2-dioxygenase, one of two isofunctional copies of the key enzyme in the gentisate pathway, and enzymes of the TCA cycle, pyruvate metabolism and gluconeogenesis were part of this group. Other proteins that are part of the sigma(54) regulon include enzymes implicated in nitrogen metabolism, transport proteins, stress-response proteins and proteins involved in cell motility. The results of this study showed that sigma(54) plays a global regulatory role in the expression of a wide variety of genes in P. alcaligenes, including the wild-type response to the presence of the aromatic inducer, gentisate.

Electrophoresis, Gel, Two-Dimensional↗

PubMed2005

Modeling depression: social dominance-submission gene expression patterns in rat neocortex.

Gene expression profiles in the cortex of adult Long-Evans rats as a function of a stressful social loss and victory in inter-male fighting encounters were examined. This social dominance and subordination model has been postulated to simulate early changes in the onset of depression in the losers. Microarrays were fabricated containing 45mer oligonucleotides spotted in quadruplicate and representing 1178 brain-associated genes. Dynamic range, discrimination power, accuracy and reproducibility were determined with standard mRNA "spiking" studies. Gene expression profiles in dominant and subordinate animals were compared using a "universal" reference design [Churchill GA (2002) Fundamentals of experimental design for cDNA microarrays. Nat Genet 32 (Suppl):490-495]. Data were analyzed by significance analysis of microarrays using rank scores [Tusher VG, Tibshirani R, Chu G (2001) Significance analysis of microarrays applied to the ionizing radiation response. Proc Natl Acad Sci USA 98:5116-5121; van de Wiel MA (2004) Significance analysis of microarrays using rank scores. Kwantitatieve Methoden 71:25-37]. Ontological analyses were then performed using the GOMiner algorithm [Zeeberg BR, Feng W, Wang G, Wang MD, Fojo AT, Sunshine M, Narasimhan S, Kane DW, Reinhold WC, Lababidi S, Bussey KJ, Riss J, Barrett JC, Weinstein JN (2003) GoMiner: a resource for biological interpretation of genomic and proteomic data. Genome Biol 4(4):R28]. And finally, genes of special interest were further studied using quantitative reverse transcriptase polymerase chain reaction. Twenty-two transcripts were statistically significantly differentially expressed in the neocortex between dominant and subordinate animals. Ontological analyses revealed that significant gene changes were clustered primarily into functional neurochemical pathways associated with protein biosynthesis and cytoskeletal dynamics. The most robust of these were the increased expression of interleukin-18, heat shock protein 27, beta3-tubulin, ribosome-associated membrane protein 4 in subordinate animals. Interleukin-18 has been found to be over-expressed in human depression and panic disorder as well as other physiological stress paradigms [Takeuchi M, Okura T, Mori T, Akita K, Ohta T, Ikeda M, Ikegami H, Kurimoto M (1999) Intracellular production of interleukin-18 in human epithelial-like cell lines is enhanced by hyperosmotic stress in vitro. Cell Tissue Res 297(3):467-473] and heat shock proteins have been shown to be involved in the pathogenesis of many neurodegenerative and psychiatric disorders [Iwamoto K, Kakiuchi C, Bundo M, Ikeda K, Kato T (2004) Molecular characterization of bipolar disorder by comparing gene expression profiles of postmortem brains of major mental disorders. Mol Psychiatry 9(4):406-416; Pongrac JL, Middleton FA, Peng L, Lewis DA, Levitt P, Mirnics K (2004) Heat shock protein 12A shows reduced expression in the prefrontal cortex of subjects with schizophrenia. Biol Psychiatry 56(12):943-950]. Thus, the gene expression changes that we have observed here are consistent with and extend the observations found in the clinical literature and link them to the animal model used here thereby reinforcing its use to better understand the genesis of depression and identify novel therapeutic targets for its treatment.

Animals↗

PubMed2005

A differential proteome in tumors suppressed by an adenovirus-based skin patch vaccine encoding human carcinoembryonic antigen.

We created an anti-tumor vaccine by using adenovirus as a vector which contains a cytomegalovirus early promoter-directed human carcinoembryonic antigen gene (AdCMV-hCEA). In an attempt to develop the skin patch vaccine, we epicutaneously vaccinated Balb/c mice with AdCMV-hCEA. After nine weeks post-immunization, vaccinated mice evoked a robust antibody titer to CEA and demonstrated the capability of suppressing in vivo growth of implanted murine mammay adenocarioma cell line (JC-hCEA) tumor cells derived from a female Balb/c mouse. Proteomic analysis of the tumor masses in the non-vaccinated naive and vaccinated mice reveal that six proteins change their abundance in the tumor mass. The levels of adenylate kinase 1, beta-enolase, creatine kinase M chain, hemoglobin beta chain and prohibitin were statistically increased whereas the level of a creatine kinase fragment, which is undocumented, was decreased in the tumor of vaccinated mice. These proteins may provide a vital link between early-stage tumor suppression and immune response of skin patch vaccination.

Adenoviridae↗

PubMed2004

The comparative analysis of serum proteomes for the discovery of biomarkers for acute myeloid leukemia.

OBJECTIVE: Acute myeloid leukemia (AML) develops as the consequence of a series of genetic changes in a hematopoietic precursor cell. However, the definitive diagnostic protein biomarkers for AML are still unclear. In our study to identify the biomarkers for an initial diagnosis, detection of relapse, and monitoring the minimal residual disease in AML by a less invasive method, serum proteins reflecting alterations in their proteomes were analyzed. MATERIALS AND METHODS: We compared the two-dimensional electrophoresis patterns of human sera of 12 patients with AML with those of 12 normal subjects. The differentially expressed spots were identified by matrix-assisted laser desorption/ionization time-of-flight and electrospray ionization quadupole time-of-flight mass spectrometries. RESULTS: Eight proteins that expressed differentially in the AML group were found. The expression levels of alpha-2-HS-glycoprotein, complement-associated protein SP-40, 40, RBP4 gene product, lipoprotein C-III, and an unknown protein were downregulated in serum of AML patients, whereas the other three proteins, including immunoglobulin heavy-chain variant, proteosome 26S ATPase subunit 1, and haptoglobin-1 were upregulated. CONCLUSION: These results suggest that these proteins can be used as less invasive diagnostic and monitoring biomarkers of AML if further studies are done.

Adult↗

PubMed2004

Quantitative analysis of liver protein expression during hibernation in the golden-mantled ground squirrel.

Mammals that enter deep hibernation experience extreme reductions in body temperature and in metabolic, respiratory, and heart rates for several weeks at a time. Survival of these extremes likely entails a highly regulated network of tissue- and time-specific gene expression patterns that remain largely unknown. To date, studies to identify differentially-expressed genes have employed a candidate gene approach or in a few cases broader unbiased screens at the RNA level. Here we use a proteomic approach to compare and identify differentially expressed liver proteins from two seasonal stages in the golden-mantled ground squirrel (summer and entrance into torpor) using two-dimensional gels followed by MS/MS. Eighty-four two-dimensional gel spots were found that quantitatively alter with the hibernation season, 68 of which gave unambiguous identifications based on similarity to sequences in the available mammalian database. Based on what is known of these proteins from prior research, they are involved in a variety of cellular processes including protein turnover, detoxification, purine biosynthesis, gluconeogenesis, lipid metabolism and mobility, ketone body formation, cell structure, and redox balance. A number of the enzymes found to change seasonally are known to be either rate-limiting or first enzymes in a metabolic pathway, indicating key roles in metabolic control. Functional roles are proposed to explain the changes seen in protein levels and their potential influence on the phenotype of hibernation.

Amino Acid Sequence↗

PubMed2005

Proteome profiling of corneal epithelium and identification of marker proteins for keratoconus, a pilot study.

The purpose of this study is to identify corneal proteins differentially expressed between keratoconus and normal epithelial samples. Proteins from the corneal epithelium were isolated from 6 keratoconus and 6 myopia patients (controls) and separated by 2D-gel electrophoresis. Six % and 12% SDS-PAGE gels were used to separate low and high molecular weight proteins. Gels were silver stained and protein spots were defined by Melanie II software. The proteins that were most altered in expression comparing keratoconus and controls were extracted, trypsin-digested, and identified by mass spectroscopy. Approximately 200-500 protein spots were detected on each gel. Nineteen spots were identified as differentially expressed between keratoconus and reference epithelium including cytokeratin 3 (< 7.8 fold), gelsolin (1.6 fold), S100A4 (1.9 fold), and enolase 1 (0.72 fold). Another identified protein found at very high levels was cytokeratin 12. Gelsolin, cytokeratin 3, and cytokeratin 12 have previously been described to be involved in other corneal diseases. Three proteins, gelsolin, alpha enolase, and S100A4 were identified to be differentially expressed in keratoconus compared to reference epithelium and thus may be involved in the pathogenesis.

Biomarkers↗

PubMed2005

A fast method for quantitative proteomics based on a combination between two-dimensional electrophoresis and 15N-metabolic labelling.

We provide a method for accurate protein quantitation that uses two-dimensional (2-D) gel electrophoresis for protein separation, but does not require extensive statistical analysis of staining intensities on gels. Instead, accurate quantitation occurs on the mass spectrometer (MAS) on multiple peptides to provide statistical evidence. In an example study, Sulfolobus solfataricus cells were grown on the carbon sources glucose, fructose and glutamate. The glucose phenotype (reference) was grown on (15)N-enriched medium. Next, the glutamate and the fructose phenotypes are mixed with the reference and two 2-D gels are created. Staining intensities of gel spots in this case are used for initial, semiquantitative assessment of differential expression. On this basis, spots are selected for accurate quantitation on the MAS. A number of differentially expressed proteins were found, for example: a (25.2 +/- 8.2)-fold upregulation of isocitrate lyase and a (7.14 +/- 0.82)-fold downregulation of glucose dehydrogenase on glutamate compared to glucose. With this protocol, intergel and interlaboratory comparisons are facilitated, since the light and heavy versions of a protein are equally affected by variations in sample preparation and buffer composition. Because the statistical evidence is gathered on the MAS, the need to run vast numbers of gels is removed.

Down-Regulation↗

PubMed2004

Effects of benzo(a)pyrene on protein expression in Jurkat T-cells.

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental pollutants of air, water and soil, and are produced by the incomplete combustion of organic materials. The International Agency for Research on Cancer has characterized PAHs as carcinogens. In this study, we investigated the effects of benzo(a)pyrene (B(a)P), which is the most carcinogenic member of the PAHs, on Jurkat cell protein by proteomic analysis. Jurkat cells were treated with various concentrations of B(a)P (0, 2.5, 5, 10, 20 or 40 microM) for 24 or 48 h and 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium and lactate dehydrogenase assays were carried out to determine cytotoxicity and a Comet assay was used to determinate genotoxicity. The cytotoxicity assays showed that 2.5 microM of B(a)P was the maximal concentration that did not cause any toxicity, but nevertheless, at this level B(a)P produced significant DNA damage in Jurkat cells at 48 h. Proteomic analysis using three different pI ranges and large two-dimensional gel electrophoresis showed 3427 protein spots. A total of 46 (13 up- and 33 down-regulated) proteins were identified as biomarkers of B(a)P and showed dose-dependent expressions in Jurkat T-cell line exposed to B(a)P. Of these, 27 protein spots were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry. Two functionally differentiated protein groups were found. The protein group involving apoptosis and tumor suppression were found to be up-regulated, and B(a)P down-regulated enzyme was involved in energy metabolism, DNA synthesis and in cell structure and motility.

Benzo(a)pyrene↗

PubMed2025

Diagnosing the undiagnosed: AI-enhanced multimodal modeling for placental mesenchymal dysplasia in high-risk pregnancies.

Placental mesenchymal dysplasia (PMD) is a rare vascular placental disorder that mimics molar pregnancy but often coexists with a viable fetus, making its misdiagnosis potentially devastating. In high-risk pregnancies, artificial intelligence (AI)-enhanced multimodal modeling - incorporating imaging, genomics, proteomics, and clinical features - offers a transformative diagnostic strategy. Leveraging Bayesian hyperparameter optimization for model refinement, this approach improves diagnostic accuracy while reducing uncertainty and clinician hesitation. Recent clinical studies support its efficacy and interpretability through SHAP and LIME models, while real-time surgical enhancements using Bayesian methods highlight its broader clinical utility. Despite current challenges such as data heterogeneity and integration barriers, multimodal AI provides unprecedented resolution in placental analysis, enabling precise differentiation between PMD and similar fetopathies. Ultimately, this advancement supports timely, non-invasive diagnosis, personalized management, and emotionally informed decision-making aligned with ethical AI implementation standards.

Bayesian optimization↗

PubMed2005

Calcium influences cellular and extracellular product formation during biofilm-associated growth of a marine Pseudoalteromonas sp.

Bacteria undergo a variety of physiological changes following a switch from planktonic growth to surface-associated biofilm growth. Here, it is shown that biofilm development of a marine isolate, Pseudoalteromonas sp. 1398, results in global changes in its cytosolic and extracellular proteomes. Calcium influences these proteome responses, and affects the amount of surface-associated biomass and extracellular matrix material produced by Pseudoalteromonas sp. 1398. Four extracellular proteins, characterized by N-terminal sequencing, showed increased abundances, while one protein, flagellin, showed reduced abundance at higher [Ca(2+)]. Immunoblotting and transmission-electron-microscopy analysis confirmed that higher [Ca(2+)] and surface-associated growth results in the repression of flagella production. Two-dimensional gel electrophoresis (2DGE) studies combined with cluster analysis of global proteome responses demonstrated that Ca(2+) had a greater regulatory influence on Pseudoalteromonas sp. growing in biofilms than on planktonic cultures. Approximately 22 % of the total cytosolic proteins resolved by 2DGE had differing abundances in response to a switch from planktonic growth to surface-associated growth when the cells were cultivated in 1 mM Ca(2+). At higher [Ca(2+)] this number increased to 38 %. Fifteen cellular proteins that were differentially expressed in response to biofilm growth and/or Ca(2+) were analysed by N-terminal sequencing and/or MS/MS. These proteins were identified as factors involved in cellular metabolic functions, putative proteases and transport proteins, although there were several proteins that had not been previously characterized. These results indicate that Ca(2+) causes global changes in matrix material, as well as in cellular and extracellular protein profiles of Pseudoalteromonas sp. 1398. These changes are more pronounced when the bacterium grows in biofilms than when it grows in planktonic culture.

Bacterial Proteins↗

PubMed2003

Model building and model checking for biochemical processes.

A central claim of computational systems biology is that, by drawing on mathematical approaches developed in the context of dynamic systems, kinetic analysis, computational theory and logic, it is possible to create powerful simulation, analysis, and reasoning tools for working biologists to decipher existing data, devise new experiments, and ultimately to understand functional properties of genomes, proteomes, cells, organs, and organisms. In this article, a novel computational tool is described that achieves many of the goals of this new discipline. The novelty of this system involves an automaton-based semantics of the temporal evolution of complex biochemical reactions starting from the representation given as a set of differential equations. The related tools also provide ability to qualitatively reason about the systems using a propositional temporal logic that can express an ordered sequence of events succinctly and unambiguously. The implementation of mathematical and computational models in the Simpathica and XSSYS systems is described briefly. Several example applications of these systems to cellular and biochemical processes are presented: the two most prominent are Leibler et al.'s repressilator (an artificial synthesized oscillatory network), and Curto- Voit-Sorribas-Cascante's purine metabolism reaction model.

Biochemical Phenomena↗