Posttranslational modifications protein modification detection

Recent comparison of gene expression profiles of von Hippel-lindau-i- versus von Hippel-lindau-ceU lines obtained from the clear cell RCC subtype identified proteins that might serve as candidate molecular markers. Inactivation of VHL is a hallmark in most sporadic clear cell RCC and it occurs early in renal carcinogenesis (Skates and Iliopoulus., 2004 Latifet al, 1993). Furthermore, the profile of secreted proteins could be directly AQ4 determined by analyzing comparatively the genomic and proteomic patterns of these cell lines as well as their conditioned tissue culture supernatants (Ferguson et al, 2004). A combination of cDNA microarray and proteome analyses appears reasonable since not every difference at the transcriptome level will translate into differences at the protein level, whereas posttranscriptional/posttranslational modifications were not detectable by tran-scriptomics. 

Size-based analysis of SDS-protein complexes in polyacrylamide gels (SDS-PAGE) is the most common type of slab gel electrophoresis for the characterization of polypeptides, and SDS-PAGE is one of the most commonly used methods for the determination of protein molecular masses.117 The uses for size-based techniques include purity determination, molecular size estimation, and identification of posttranslational modifications.118119 Some native protein studies also benefit from size-based separation, e.g., detection of physically interacting oligomers. 

Carr SA, Annan RS, Huddleston MJ. Mapping posttranslational modifications of proteins by MS-based selective detection Application to phosphoproteomics. In Burlingame AL, ed.. Mass Spectrometry Modified Proteins and Glycoconjugates, Vol. 405, New York Academic Press, 2005, 82-115. 

After the early discovery of a tyrosine 0-sulfate residue in bovine fibrinopeptide B, 15 this posttranslational modification which occurs ubiquitously in proteins was also detected in a series of biologically active peptides such as the neurohormones of the gastrin/cholecysto-kinin (CCK) family of peptides, phyllokinin, Leu-enkephalin, and the thrombin inhibitor hirudin listed in Table 1. 

One problem associated with the yeast 2-hybrid system is that protein interactions that are dependent on posttranslational modifications that can occur only in mammalian cells, would not be detected. For example, monoubiquitinylation and serine phosphorylation of a Fanconi anemia complementation group D2 protein (FANC-D2) are both required for mediating cellular resistance to ionizing radiation (122), an interaction that would be missed using this approach. 

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