Proteins: Analysis and Design
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Proteins: Analysis and Design focuses solely on individual experimental approaches, rather than on specific classes of proteins. The book provides insight into the important issues in protein science and how one can cope with them. These include all issues which explore the detailed relationship of protein structure to function.
* Provides problems and technical solutions
* Includes posttranslational modifications
* Uses synthetic peptides as biological models
* Details mutagenesis and protein engineering
* Covers design of protein structure and function
or digestion. 34 Ruedi Aebersold and Scott D. Patterson 2. Peptide-Mass Mapping The size of the contents of genomic and cDNA sequence databases is growing exponentially. The significance of de novo amino acid sequence determination for the purpose of cloning the coding gene is therefore diminishing, at least for those species that are the primary focus of genome sequencing projects. Concurrently, the need for rapid, sensitive, and conclusive methods of identifying the gene coding for a
calculating all the possible peptide masses that can be generated by fragmenting all the sequences in a sequence database under certain, deliberately chosen parameters [(280,281,297-299), and reviewed in (300,301)]. All of these programs are available as commercial packages, from the authors, across the Internet as an e-mail server [email@example.com, (281)], or by accessing the Darwin suite of programs on the World Wide Web (URL http://cbrg.inf.ethz.ch/) (241,297). Other programs that offer similar
data. (iv) The molecular mass suggests the origin of the fragment in proteolytic digests. Proteolytic digestion of small amounts of peptides requires relatively high enzyme-to-substrate ratios. Therefore, some of the isolated peptides may be derived from autolysis of the protease. Comparison of the determined peptide masses with the peptide masses predicted from autolysis identifies such peptides. (v) The peptide mass can be used to complete a peptide sequence in cases in which a residue could
monitoring the tyrosine phosphorylation status of proteins separated by 1-D or 2-D gel electrophoresis. It should be noted, however, that lower molecular weight tyrosine phosphorylated proteins may not be detected as efficiently (413,414) as proteins with larger molecular weight. Numerous groups have attempted to produce antisera with specificity for phosphothreonine or phosphoserine. A polyclonal antiserum with specificity for phosphothreonine was reported (415), and Sigma Chemical Company (St.
protein complex forms the transcription factor that activates immediate-early gene transcription. By convention, the latent cytoplasmic transcription factors have been termed STATs (signal transducers and activators of transcription), whereby the 91and 84-kDa proteins are named Stat lc~ and Stat 113, respectively, and the 113kDa polypeptide is named Stat 2. In this chapter we have chosen this pathway (shown in Fig. 1), which has been described in more detail in several excellent reviews (27-29),