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Data and increases the level of confident matches. Anther protein identification is typically carried out by N-terminal Edman sequencing on the Procise 494-01 sequencer system (Perkin-Elmer Life Sciences, Boston, MA) and by MS on Micromass TofSpec 2E-TOF-MS (Manchester, UK) or PE SCIEX (Foster City, CA), QSTAR hybrid LC-MS MS Q-TOF, or Micromass LC-MS MS Q-TOF systems. Search engines such as Profound, MassLynx, and Mascot work well for both MS data and MS MS data. However, a key component of the searches is the databases. Besides general nonredundant database, species-specific EST databases or any other specific databases are needed for high success rates in protein identification. A flow chart for the analysis of anther proteomes by 2-DGE MS approach is shown in Figure 17.3.
The anther is by far the best-characterized by proteomics 20 , for which high-quality 2D gel reference maps have been generated during male gametophyte development in the rice cultivar Doongara under strictly controlled environmental conditions 15, 16 . Anthers at the young microspore stage (from tetrad stage to early microspore stage) were separated by 2-DGE, and silver staining detected over 4000 spots in the pH range of 4-11 that represent approximately 10 of the estimated total genomic output of rice. Qualitative and quantitative analysis of differentially displayed spots was performed with Melanie 3 software. Two-hundred and seventy-three spots, collected from either PVDF membranes or colloidal CBB-stained 2D gels, were analyzed by Edman sequencing and MALDI-TOF-MS or Q-TOF-LC-MS MS. MS analysis was performed with the search engine Profound and MassLynx to search nonredundant database and rice EST database. A total of 53 spots were identified that represented 43 different...
Most of the proteins that have been shown to bind MTs in plant cells are classical MAPs that have roles in regulating the dynamics and organization of MTs. Others fall into the nonclassical MAP category. In Table 19.1 we have listed examples of plant-specific proteins that have authentic MT-binding activity in situ, as well as proteins that are common to eukaryotes but have been shown to localize to MTs in plant cells only. Numerous other predicted MT-binding proteins have been identified using biochemical purification techniques or using genome database search engines, but have not yet been shown to localize to MTs in situ. In this section, we discuss the identity and roles of several of these confirmed and predicted MT-binding proteins. For more complete lists, readers are directed to reviews and research publications that provide detailed lists of these proteins 4, 8, 9, 22, 35 .
Web Pages about Echinacea Found via Major Search Engines, August 2001 and 2002 Search Increase ( ) Pages Year Alta Vista related to Echinacea found when using some search engines has reached 100,000 to 450,000 per year (Table 9.3). It can be seen that the popularity of Echinacea is dramatically increasing along with the rising popularity of CAM worldwide.
While searching the Internet with three of the most popular search engines, we encountered duplicated Web pages and advertising. However, by carefully setting the search terms, it is possible to find a large quantity of useful and scientific information. Among these search engines, Google is a satisfactory one in obtaining valuable information on Echinacea. The number of Web pages
Powerful new technologies such as automated separation techniques, high-throughput screening and combinatorial chemistry are revolutionizing drug discovery. Traditional knowledge can serve as a powerful search engine, which will greatly facilitate and rediscover intentional, focused and safe natural product drug discovery. By looking at the historical trends in drug and medical developments, it is possible to understand how current drug development will benefit from this partnership.
Peptide fragmentation identification requires MS MS and allows the unambiguous identification of proteins, even from complex protein mixtures. Thus, this analysis of peptides requires the fragmentation of peptides into different ions by CID. The pattern of the fragment ions can be interpreted by database searches, since the fragmentation spectrum of a given peptide is somehow predictable (Figure 3.2). Databases predict that the fragment ions of all available peptides in the database and the theoretical and measured, ion pattern are compared to identify matches. Thus, not only the peptide mass is measured, but also amino acid sequence information is provided. The correct peptide interpretation is based on MS MS search algorithms that are provided by search engines such as Mascot 16 , SEQUEST 17 , and Phenyx 18, 19 .
The identification of proteins by MS MS is based on the correlation of MS data with database sequences. Using a search engine, fragmentation patterns of enzy-matically generated peptides are compared to theoretical data generated from protein sequence databases in silico 17 . The ever-increasing rate at which mass spectra can be accumulated means that greater reliance is placed on computer-based software to process spectra and interpret results. Search engines currently available differ in their sensitivity and in their ability to distinguish correct and incorrect peptide hits 18 . The number of incorrect identifications can be decreased by analyzing all spectra with multiple search engines such as Mascot 19 , Sequest 20 , and X Tandem 21 and then validating the combined results using Peptide Prophet 22 and Protein Prophet 23 as implemented in Scaffold (http www.proteomesoftware.com ). Different algorithmic approaches are being developed to improve protein identification searches,...
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