Current Proteomics

ISSN: 1570-1646


OPEN ACCESS PLUS


Contents


Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC) – an Introduction for Biologists, 2011, 8, 2-16
Robert L.J. Graham, Michael J. Sweredoski and Sonja Hess
[Abstract] [Full Text Article]


The Impact of Proteomic Advances on Bacterial Gene Annotation, 2009, 6, 84-92
Gustavo A. de Souza and Harald G. Wiker
[Abstract] [Full Text Article]


Deciphering the Antibodyome - Peptide Arrays for Serum Antibody Biomarker Diagnostics, 2009, 6, 1-12
H. Andresen and C. Grötzinger
[Abstract] [Full Text Article]


Recent Advances in Controlled Immobilization of Proteins onto the Surface of the Solid Substrate and Its Possible Application to Proteomics, 2008, 5, 161-175
K. Nakanishi, T. Sakiyama, Y. Kumada, K. Imamura and H. Imanaka
[Abstract] [Full Text Article]



Abstracts


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Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC) – an Introduction for Biologists
Robert L.J. Graham, Michael J. Sweredoski and Sonja Hess

[Full Text Article]

Since its introduction in 2002 ‘stable isotope labeling by amino acids in cell culture’ (SILAC) has become a major tool for quantitative proteomics. Stable isotopes are incorporated into proteins by introduction of labeled amino acids in growth medium. This methodology’s compatibility with virtually all cell culture conditions, ease of implementation into existing workflows and the high quality quantitative data that can be obtained have made it the quantitative method of choice for many laboratories. Although originally used for mammalian cell cultures, it has been adapted to a number of organisms including yeast, bacteria, plants and higher organisms. This review will look at the use of SILAC as a key tool in quantitative biology. The purpose of this review is to furnish the reader with a basic understanding of the fundamental principles of SILAC including its implementation, application, bioinformatic tools for its analyses and potential problems that one should be mindful of.


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The Impact of Proteomic Advances on Bacterial Gene Annotation
Gustavo A. de Souza and Harald G. Wiker

[Full Text Article]

Over the past decade the large use of genomic approaches has resulted in the impressive generation of complete genomic sequences for over 800 bacterial species. However, the use of different bioinformatic approaches to determine the presence of a gene or open reading frame (ORF) in those genomes cause divergent gene annotations, even for data generated from the same genomic sequences. The use of a correct dataset for protein identification is a key step in many fields as phylogenetics, protein expression experiments, and has an impact on the identification capacity of a proteomic workflow. In this review, we describe successful attempts performed by proteomic groups to improve gene annotation in bacteria using different bioinformatic and mass spectrometry technologies. The review emphasizes the most recent advances in high resolution MS technology, which has increased the sequence coverage and peptide identification reliability by several fold. The capacity to perform deeper and more complete catalogations allows correction of several known genes, plus the discovery of protein products of regions of the genome not yet predicted to be coding areas. Recent results from our group show how such technology can be used as a guide to correct mistakes in transcriptional starting site (TSS) choices of proteins of Mycobacterium tuberculosis and Mycobacterium leprae, as well as to identify N-terminal peptides resulting from signal peptidase cleavages.


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Deciphering the Antibodyome - Peptide Arrays for Serum Antibody Biomarker Diagnostics
H. Andresen and C. Grötzinger

[Full Text Article]

The analysis of antibodies in human serum is an established technique in the laboratory diagnosis of infectious as well as autoimmune diseases. The multitude of antibody reactions towards pathogens and likewise the antibody profile in autoimmune diseases does contain a wealth of proteomic (antibody) data that may constitute valuable diagnostic information with relevance for the patient’s prognosis and response to therapy. Hence the use of antibodies as diagnostic biomarkers may be one of the most promising strategies to identify patient subgroups. The presence or absence of antibodies directed against specific epitopes could represent a serologic biomarker that is able to predict the severity of a disease and assist in medical decision making. In addition, parallel detection of many different antibodies in a serum sample would be of great value in many areas of basic immunological research. Peptide arrays displaying biologically active small synthetic peptides in either low, medium or high-density formats represent an attractive technology to probe complex serum samples for the presence of such antibody analytes. Holding the unique capacity to break down the heterogeneous immunologic response into monoclonal antibody specificities and to differentiate subtle changes in antibody abundance and specificity, the peptide array technology by far extends the diagnostic potential of any conventional serologic assay. Together with an unrivalled parallelity, peptide (micro)array analysis opens new perspectives for the novel use of antibodies as diagnostic biomarkers and provides unique access to a more differentiated serological diagnosis. This review recapitulates the development of the peptide array technology with a focus on recent advances and current state of the art platforms for antibody diagnostics. Latest applications of peptide arrays for the serologic diagnosis of infectious diseases, autoimmunity and allergy are discussed, and conclusions for future developments and implications are drawn.


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Recent Advances in Controlled Immobilization of Proteins onto the Surface of the Solid Substrate and Its Possible Application to Proteomics
K. Nakanishi, T. Sakiyama, Y. Kumada, K. Imamura and H. Imanaka

[Full Text Article]

Proteome analysis plays a key role in the elucidation of the functions and applications for numerous proteins. For proteome analyses, various microplate- and microarray-based techniques have been developed by a number of researchers. Their intent was to immobilize proteins on the surface of a solid substrate in a site-directed manner while retaining structure and native biological function. In this review, we focus on recent advances in immobilization methodology for proteins/enzymes on a surface, including those using the affinity peptides screened by random peptide library systems. We also discuss applications of the affinity peptide-mediated immobilization method in fields related to proteome analysis, particularly our recent work concerning immunoassay and protein-protein interaction analysis.