Posttranslational modifications tools

Meri, S. andBaumann, M., Proteomics posttranslational modifications, immune responses and current analytical tools, Biomol. Eng., 18, 213, 2001. 

ExPASy Proteomics tools (http //expasy.org/tools/), tools and online programs for protein identification and characterization, similarity searches, pattern and profile searches, posttranslational modification prediction, topology prediction, primary structure analysis, or secondary and tertiary structure prediction. 

Pathological conditions are also linked to posttranslational modifications such as oxidized histidine residues found in P-amyloid protein of Alzheimer s patients, or conformational variants in the case of prion-induced encephalopathies. The development of sensitive MS tools and proteomics techniques is playing an active role in the precise description of these mechanisms.97,98... 

Figure 3. Role of nonhistone protein acetylation in maintaining cellular homeostasis- mis-regulation and disease connection (a) Acetylation of nonhistone proteins are associated with active or repressed chromatin architecture as guided by suitable cellular signals for maintenance of gene expression. Misreg-ulation of HAT function leads to diseased state, where chromatin architecture is altered than under normal condition. In a parallel way the posttranslational modification status of these proteins may act as versatile tool to diagnose the various stages of disease manifestation e.g. probable involvement of acetylated NPMl modulating its stress response function can lead us to use it as a marker for various disease states, (b) Acetylation of nonhistone proteins in connection to diseases like Cancer, AIDS, Diabetes and others. (See Colour Plate 14.)...

How do we then envision the protein microarray as a proteomics tool We now estimate the human genome to comprise around 30,000 genes. For gene expression analysis using DNA microarrays, 1000 to 10,000 gene elements are often used. Since proteins undergo posttranslational modification (>200 different types see McDonald and Yates, 2000, Reference 40) and can occur as isoforms and multiprotein complexes, the number of protein expression elements needs to be much larger. 

Isoelectric focusing is a mature separation technique that has a place in any laboratory doing work with proteins. The analysis of a protein is not complete without a determination of its isoelectric point and all protein databases have at least estimates of the pis of the represented proteins. Proteins thought to be pure by other methods are often found to be mixtures of several proteins when analyzed by IEF. Isomeric forms of the same protein that are revealed by IEF are valuable indicators of mutations or differences in posttranslational modifications. IEF plays a crucial role in 2-D PAGE and preparative IEF allows for high-purity fractionations of unparalleled resolution. A simple keyword search of literature databases shows that about 500 journal articles are written per year referring to IEF. This amply attests to the value of IEF as a tool for protein analysis and purification. 

Understanding the action mechanism of flavoenzymes heavily relies on the combination of different chemical tools and techniques. First, it is of utmost importance to have a pure and stable (recombinant) protein. Size exclusion chromatography will provide information about the enzyme quaternary structure, and mass spectrometry can establish posttranslational modifications. For a detailed insight into the protein structure, well-diffracting crystals are needed to determine the X-ray structure. 

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