To demonstrate the fructose metabolism pathway in Bifidobacterium Longum NCC2705 and to construct its fermentation model, we explored the comparative proteome cultivating the strain on glucose or fructose, based on a proteomic reference map of B. longum NCC2705 constructed earlier. Then, we used matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry and electro-spray ionization tandem mass spectrometry (ESI-MS/MS) for differently expressed proteins identification. Furthermore, with semi-quantitative RT-PCR we determined the distinctively expressed proteins at the level of transcription. Proteomic comparison of glucose- and fructose-grown cells demonstrated much similarity. On the page of fructose there were all the enzymes and proteins that exist during the process of glucose degradation. We observed a greater variation of more than three-fold for the identified 9 spots representing 5 protein entries by MALDI-TOF MS. The sugar-binding protein specific to fructose (BL0033) and an ABC transporter ATP binding protein (BL0034) showed higher expression level from cells grown on fructose. It was also determined by semi-quantitative RT-PCR subsequently. BL0033 time course and concentration experiments showed that the induction time correlated to higher fructose concentration, and increased expression of BL0033. Fructose was catabolized via the same degradation pathway as glucose at the level of proteomics. BL0033 was induced by fructose. All results suggest that the uptake of fructose into the cell may be conducted by a specific ABC transport system, in which BL0033 and BL0034 as components might have played an important role.
Bifidobacterium ; chemistry ; genetics ; metabolism ; Culture Media ; Fermentation ; Fructose ; pharmacology ; Glucose ; pharmacology ; Proteome ; analysis ; genetics ; Proteomics ; methods

In this study, we investigated the mechanism of transformation by Bacillus subtilis strain 168 by proteomic analysis. B. subtilis strain 168 was able to stereoselectively transform cis-propenylphosphonic acid (cPPA) to fosfomycin. The maximal fosfomycin production was 816.6 microg/mL after two days cultivation, with a conversion rate of 36.05%. We separated the whole cellular proteins by two-dimensional gel electrophoresis (2-DE) method, and 562 protein spots were detected in the presence of cPPA in the medium, while 527 protein spots were detected in the absence of cPPA. Of them, 98 differentially expressed protein spots were found. Among them, 52 proteins were up-regulated whereas 20 were down-regulated in the presence of cPPA in the medium, and 26 induced at the presence of cPPA. The differentially expressed proteins were analyzed by combined MS and MS/MS methods. Eighty protein spots, including 45 up-regulated proteins, 17 down-regulated proteins, and 18 induced by cPPA were identified. Based on the results of proteomic analysis, we postulated two steps of transformation: in the first step, cPPA was hydrated to 2-hydroxypropylphosphonic acid; in the second step, 2-hydroxypropylphosphonic acid was transformed to fosfomycin via a dehydrogenation reaction.
Bacillus subtilis ; genetics ; growth & development ; metabolism ; Bacterial Proteins ; metabolism ; Biotransformation ; Fosfomycin ; metabolism ; Organophosphorus Compounds ; metabolism ; Proteome ; metabolism ; Proteomics

The prevalence of obesity has increased worldwide in recent decades. Genetic factors are now known to play a substantial role in the predisposition to obesity and may contribute up to 70% of the risk for obesity. Technological advancements during the last decades have allowed the identification of many hundreds of genetic markers associated with obesity. However, the transformation of current genetic variant-obesity associations into biological knowledge has been proven challenging. Genomics and proteomics are complementary fields, as proteomics extends functional analyses. Integrating genomic and proteomic data can help to bridge a gap in knowledge regarding genetic variant-obesity associations and to identify new drug targets for the treatment of obesity. We provide an overview of the published papers on the integrated analysis of proteomic and genomic data in obesity and summarize four mainstream strategies: overlap, colocalization, Mendelian randomization, and proteome-wide association studies. The integrated analyses identified many obesity-associated proteins, such as leptin, follistatin, and adenylate cyclase 3. Despite great progress, integrative studies focusing on obesity are still limited. There is an increased demand for large prospective cohort studies to identify and validate findings, and further apply these findings to the prevention, intervention, and treatment of obesity. In addition, we also discuss several other potential integration methods.
Humans ; Proteome/metabolism* ; Proteomics ; Prospective Studies ; Obesity/genetics* ; Genomics ; Genome-Wide Association Study

Cells sense various in vivo mechanical stimuli, which initiate downstream signaling to mechanical forces. While a body of evidences is presented on the impact of limited mechanical regulators in past decades, the mechanisms how biomechanical responses globally affect cell function need to be addressed. Complexity and diversity of in vivo mechanical clues present distinct patterns of shear flow, tensile stretch, or mechanical compression with various parametric combination of its magnitude, duration, or frequency. Thus, it is required to understand, from the viewpoint of mechanobiology, what mechanical features of cells are, why mechanical properties are different among distinct cell types, and how forces are transduced to downstream biochemical signals. Meanwhile, those in vitro isolated mechanical stimuli are usually coupled together in vivo, suggesting that the different factors that are in effect individually could be canceled out or orchestrated with each other. Evidently, omics analysis, a powerful tool in the field of system biology, is advantageous to combine with mechanobiology and then to map the full-set of mechanically sensitive proteins and transcripts encoded by its genome. This new emerging field, namely mechanomics, makes it possible to elucidate the global responses under systematically-varied mechanical stimuli. This review discusses the current advances in the related fields of mechanomics and elaborates how cells sense external forces and activate the biological responses.
Biomechanical Phenomena ; Gene Expression Regulation ; Humans ; Mechanotransduction, Cellular ; Models, Biological ; Proteome ; genetics ; metabolism ; Stress, Physiological ; Transcriptome

This study is aimed at gaining insights into the brain site-specific proteomic senescence signature while comparing physiologically aged brains with aging-related dementia brains (for example, Alzheimer's disease (AD)). Our study of proteomic differences within the hippocampus (Hp), parietal cortex (pCx) and cerebellum (Cb) could provide conceptual insights into the molecular mechanisms involved in aging-related neurodegeneration. Using an isobaric tag for relative and absolute quantitation (iTRAQ)-based two-dimensional liquid chromatography coupled with tandem mass spectrometry (2D-LC-MS/MS) brain site-specific proteomic strategy, we identified 950 proteins in the Hp, pCx and Cb of AD brains. Of these proteins, 31 were significantly altered. Most of the differentially regulated proteins are involved in molecular transport, nervous system development, synaptic plasticity and apoptosis. Particularly, proteins such as Gelsolin (GSN), Tenascin-R (TNR) and AHNAK could potentially act as novel biomarkers of aging-related neurodegeneration. Importantly, our Ingenuity Pathway Analysis (IPA)-based network analysis further revealed ubiquitin C (UBC) as a pivotal protein to interact with diverse AD-associated pathophysiological molecular factors and suggests the reduced ubiquitin proteasome degradation system (UPS) as one of the causative factors of AD.
Aged, 80 and over ; Alzheimer Disease/*metabolism ; Brain/*metabolism ; Female ; Gelsolin/genetics/metabolism ; Humans ; Male ; Membrane Proteins/genetics/metabolism ; Neoplasm Proteins/genetics/metabolism ; Organ Specificity ; Proteome/genetics/*metabolism ; Tenascin/genetics/metabolism ; Ubiquitin C/genetics/metabolism

Protein phosphorylation in bacteria is important for signaling and metabolic activity. Clostridium acetobutyicum can synthesize high yield of organic solvent under acidic condition. How solventogenesis is regulated at molecular level in this bacterium, is not clearly elucidated yet. We used two dimensional electrophoresis (2-DE) and mass spectrometry to have a differential analysis of the bacterial proteins from Clostridium acetobutylicum at acedogenic and solventogenic stage. We focused on these iso-spots with similar molecular mass and different pI values. Totally, eight string spots were identified, which displayed significant changes of pI values as well as spot volumes in response to solventogenic development. The data acquired from mass spectrometry demonstrated that all of the iso-spots contained the phosphrylated peptides. Bioinformatic analysis revealed that these proteins partake in the pathways of solvent synthesis.
Bacterial Proteins ; metabolism ; Clostridium acetobutylicum ; genetics ; metabolism ; Electrophoresis, Gel, Two-Dimensional ; Mass Spectrometry ; Phosphorylation ; Proteome ; analysis ; Proteomics ; methods

AIMTo search for and application of new method for data processing in metabonomic studies.

METHODSThe paper proposed that in the processing of metabonomic data, robust PCA method can be used to diagnose outliers; and unstable variables judged by comparison between difference within class and difference among classes should be excluded before data analysis; moreover, the data should be properly scaled before further processing. The proposed methods were used to preprocess metabolomic data of four genotypes of the Arabidopsis thaliana plants.

RESULTS AND CONCLUSIONThe outcome demonstrated that the application of these methods can obviously improve clustering and biomarker identifying results.

Algorithms ; Arabidopsis ; genetics ; metabolism ; Arabidopsis Proteins ; genetics ; metabolism ; Automatic Data Processing ; Gene Expression Profiling ; Gene Expression Regulation, Plant ; Genotype ; Metabolism ; Principal Component Analysis ; Proteome ; metabolism ; Proteomics ; methods

OBJECTIVETo predict the exported proteins of the novel bacterium Phenylobacterium zucineum HLK1(T) using genome-wide computational identification by searching the export signals including N-terminal signal peptides and alpha-transmembrane helices.

METHODSThe computational identification of signal sequences was based on a consensus between multiple predictive tools, including SignalP V3.0, LipoP V1.0, Phobius and TMHMM 2.0. Type IV signal peptides and proteins exported via TAT machinery were searched manually based on the conservative motifs. All the predicted proteins were classified according to the Cluster of Orthologous Group (COG) standard.

RESULTIn the total 3861 proteins encoded by P. zucineum HLK1(T) 1 378 (35.7%) were predicted to be exported proteins, most of which (totally 735, 19.0% of the proteome and 53.3% of all the exported proteins) were uncleavable transmembrane helices. In addition, 499 type I signal peptides (12.9%, 36.2%), 101 lipoproteins (2.6%, 7.3%) were also identified. Four Type IV signal peptides and 12 TAT proteins were detected as well. According to the COG classification standard, most of these exported proteins were P proteins related to inorganic ion transport and metabolism and S proteins whose functions were unknown.

CONCLUSIONThe genome of HLK1(T) coded various types of exported proteins which may play an important role in the interaction between P. zucineum and the host cell, and facilitate the strain to invade into the cell.

Amino Acid Sequence ; Bacterial Proteins ; genetics ; metabolism ; Caulobacteraceae ; genetics ; metabolism ; Computational Biology ; methods ; Consensus Sequence ; genetics ; Genome, Bacterial ; Molecular Sequence Data ; Protein Sorting Signals ; genetics ; Protein Transport ; genetics ; Proteome ; metabolism

This study investigated the molecular markers of DS-1-47, a component of an implantation- promoting traditional Chinese medicine consisting of Astragalus mongholicus, Atractylodes macrocephala, Scutellaria baicalensis and Dipsacales, in an attempt to clarify the molecular mechanism and action targets of DS-1-47. Controlled ovarian stimulation (COS) method was used to establish the implantation dysfunction models of mice. Animals were divided into normal pregnant group, COS model group and DS-1-47 group. Laser capture microdissection-double dimensional electrophoresis-mass spectrum (LCM-DE-MS) was used to analyze the uterine protein molecules that were possibly involved in the promotion of implantation. Twenty-three proteins in DS-1-47 group were significantly changed as compared to those in COS model group, with 7 proteins down-regulated and 16 proteins up-regulated. Except for some constituent proteins, the down-regulated proteins included collagen α-1 (VI) chain, keratin 7, keratin 14, myosin regulatory light chain 12B, myosin light polypeptide 9, heat shock protein β-7, and C-U-editing enzyme APOBEC-2; the up-regulated proteins included apolipoprotein A-I, calcium regulated protein-3, proliferating cell nuclear antigen, L-xylulose reductase, and calcium binding protein. These 23 proteins that were regulated by DS-1-47 represented a broad diversity of molecule functions. The down-regulated proteins were associated with stress and immune response, and those up-regulated proteins were related to proliferation. It was suggested that these proteins were important in regulating the uterine environment for the blastocyst implantation. By identification of DS-1-47 markers, proteomic analysis coupled with functional assays is demonstrated to be a promising approach to better understand the molecular mechanism of traditional Chinese medicine.
Animals ; Drugs, Chinese Herbal ; pharmacology ; Embryo Implantation ; drug effects ; Female ; Mice ; Ovulation Induction ; Pregnancy ; Proteome ; genetics ; metabolism ; Uterus ; drug effects ; metabolism ; physiology

OBJECTIVEOur previous investigation demonstrated that angiotensin II could induce proliferation and differentiation of hepatic stellate cells, and also could up-regulate its extracellular matrix synthesis. The objective of this study was to determine the effects of 10(-5) mol/L angiotensin II on gene expression of hepatic stellate cells.

METHODSAfter incubation with 10(-5) mol/L angiotensin II for 48 hours, the cultured hepatic stellate cells were collected. The mRNA and total protein were obtained from cell lysate and then suppression subtractive hybridization (SSH), 2D-gel electrophoresis and MALDI-TOF mass spectrometry were used to identify these cDNAs and proteins.

RESULTSA total of 36 clones from the subtracted cDNA library were sequenced and compared to sequences in the GenBank using BLAST. Of the 36 differentially expressed gene fragments from the subtracted library, 13 differentially expressed gene fragments showed significant homology to other known proteins, such as ribosomal protein, beta-actin FE-3, leucyl-tRNA synthetase, CD147, pyruvate kinase, peroxiredoxin 1, and BAT3, while 2 other gene fragments encoding protein BC097361 and BC057380 and their functions were not disclosed. About 1110 and 1008 protein spots were detected by employing the ImageMaster 2D Platinum 4.9 proteome image analysis system in angiotensin-treated hepatic stellate cells and control cells separately. Among these spots 108 proteins were up-regulated while the other 153 proteins were down-regulated. Several up-regulated proteins were chosen to be excised and in-gel digest MALDI-TOF-MS and Database analysis showed that among the high expression proteins, there were prohibitin, RBL-NDP kinase 1.8x10(4) subunit, electron transfer flavoprotein alpha-subunit, guanine nucleotide binding protein, alpha 15, and heat shock 7.0x10(4) protein 5.

CONCLUSIONOur results suggest that the up-regulation of hepatic stellate cell mRNA influences proliferation, differentiation and apoptosis of those cells. The proteins of signal transduction, metabolizing regulation, apoptosis suppression, and fibrogenesis regulation of hepatic stellate cells were up-regulated.

Angiotensin II ; pharmacology ; Apoptosis ; Cell Differentiation ; Cell Line ; drug effects ; Cell Proliferation ; Gene Expression ; Gene Library ; Hepatic Stellate Cells ; drug effects ; metabolism ; Humans ; Proteome ; metabolism ; RNA, Messenger ; genetics