Protein database search engine

Enter the accurate peptide monoisotopic mass assignments into a protein database search engine. 

FIGURE 4.12 A flow chart showing the basic steps used to identify a protein. The protein is excised from a 2-D electrophoresis gel, the gel diced, and the stain is washed. The protein is reduced and digested in the gel. The newly formed peptides are extracted and analyzed using a MS and/or MS-MS scan mode. The resulting MS and MS-MS data are entered into a protein database search engine that performs the match. 

FIGURE 4.13 A MALDI-TOF mass spectrum acquired in reflectron mode showing the tryptic peptides from a digestion of bovine serum albumin. The m/z of the monoisotopic ions are assigned and can be entered into a protein database search engine to match to the theoretical trypsin digest of BSA. 

Besides sensitive methods for the analysis of proteins, bioinformatics is one of the key components of proteome research. This includes software to monitor and quantify the separation of complex samples, e.g., to analyze 2DE images. Web-based database search engines are available to compare experimentally measured peptide masses or sequence ions of protein digests with theoretical values of peptides derived from protein sequences. Websites for database searching with mass spectrometric data may be found at http //www.expasy.ch/tools, http //prospector.ucsf. edu/ and http //www.matrixscience.com. 

SwissProt is a relatively small but highly annotated database with minimal redundancy. NCBInr and MSDB are much larger, but have multiple entries for many proteins. Other search engines may have additional database choices, and in-house copies allow custom databases to be used. 

The NCBI s Entrez is apowerful database search engine. This integrated search engine provides a menu (http //www.ncbi.nih.gov/Entrez/index.html) offering the user selections to search biomedical literature (PubMed), databases on OMIM, nucleotide sequence, protein sequence, whole genome sequences, 3D macromolecular structures, taxonomy (organisms in GenBank), SNP,... 

Peptide microsequencing has proved to be a powerful approach to identify protein components of mixtures,53-55,61 and has been applied to extracted proteins as a method to identify biological threats.56,83-85 Commercial and internet accessible search engines and databases provide adequate bioinformatics support to use sequence information produced by the mass spectrometer (e.g., http //www.matrixscience.com http //prospector.ucsf.edu/ ucsfhtml4.0/mstagfd.htm http //prowl.rockefeller.edu/). 

Mascot (http //www.matrixscience.com/), a search engine that uses mass spectrometry data to identify peptides and proteins from primary sequence databases (MSDB, SwissProt, and others). 

Most PMF search engines allow a mass for the protein to be entered the search will then consider only database entries within a window around this mass. This can be dangerous, as proteins are subject to processing/degradation, which can significantly increase or decrease their molecular mass. Mascot uses a more complex method of applying this parameter, but in most cases this can be left blank. 

An Internet link to protein sequence databases and search engines is found at www.sdsc.edu/ResTools/biotools/biotoolsl9.html and a link to Internet resources for sequence analysis is www.sdsc.edu/ResTools/biotools/biotoolsl.html. 

Enzyme and Protein Databases. Large databases of protein structure have been assembled to collate data from various laboratories around the world. The National Library of Medicine maintains a website, PubMed, which catalogues the medical literature, and a search engine that allows you to search for reference articles by topic or author... 

Links. PDB s search engine, the Structme Explorer, can be used to retrieve PDB records, as shown in Figure 5.2. The Structure Explorer is also the primary database of links to third-party aimotation of PDB structure data. There are a number of links maintained in the Structure Explorer to Internet-based three-dimensional structure services on other Web sites. Figme 5.2 shows the Structure Summary for the protein bamase (IBNR Bycroft et al., 1991). The Structure Explorer also provides links to special project databases maintained by researchers interested in related topics, such as structural evolution (FSSP Holm and Sander, 1993), structure-structure similarity (DALI Holm and Sander, 1996), and protein motions (Gerstein et al., 1994). Links to visualization tool-ready versions of the structure are provided, as well as authored two-dimensional images that can be very helpful to see how to orient a three-dimensional structure for best viewing of certain features such as binding sites. 

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