Proteins, post-translational modification

Wilkins MR et al High-throughput mass spectrometric discovery of protein post-translational modifications. J Mol Biol 1999 289 645. 

Immunoprecipitation followed by SDS-PAGE and immunoblotting, is routinely used in a variety of applications to determine the molecular weights of protein antigens, to study protein-protein interactions, to determine specific enzymatic activity, to monitor protein post-translational modifications and to determine the presence and quantity of proteins. The IP technique also enables the detection of rare proteins which otherwise would be difficult to detect since they can be concentrated up to... 

MR Wilkins, E Gasteiger, AA Gooley, BR Herbert, MP Molloy, P-A Binz, K Ou, J-C Sanchez, A Bairoch, KL Williams, DF Hochstrasser. High-throughput mass spectrometric discovery of protein post-translational modifications. J Mol Biol 289 645-657, 1999. 

Perez-Vilar J., Mabolo R. McVaugh C.T., Bertozzi C.R. and Boucher R.C. (2006). Mucin granule intraluminal organization in living mucous/goblet cells. Roles of protein post-translational modifications and secretion. J Biol Chem. 281, 4844-4855 Perez-Vilar J. (2007). Mucin granule intraluminal organization. Am J Respir Cell Mol Biol,... 

Proteomics is complicated by the fact that the absolute quantification does not always reflect the molecular function of proteins, because protein activities are highly regulated post-translationally. Post-translational modifications modulate the function of proteins and thus directly impact their capacity to participate in cellular regulatory events (Cravatt and Sorensen 2000). Due to post-translational modifications, the numbers of proteins in human are estimated to be at least three times the amount of genes. Therefore, the elucidation of protein post-translational modifications is the most important justification for biochemical and structural relationships. Hence, these modifications need to be evaluated. 

Reichner ). S, Chin, Y. E. (2004). Antibodies Immobilized as arrays to profile protein post-translational modifications in mammalian cells. 

Species differences Novel recombinant proteins Post-translational modifications Lack of modification Incorrect modification Physical structure alterations Denaturation Aggregation... 

Parallel to the efforts of identifying substrate proteins for a particular enzyme, the activity-based small molecule probes pioneered by Cravatt and coworkers can facilitate the identification of the biological functions of an enzyme that catalyzes protein post-translational modifications (145). The major advantage of this type of probes is that potentially they can detect enzymes that are in the active states, and thus can provide snapshots of enzymes that are in the active states at different development stages or different types of cells. Among enzymes that catalyze PTM, so far probes have been developed for studying proteases (145, 146), kinases (147), pTyr phosphatases (148), and protein Arg deiminases (149). 

Mapping of protein (post-translational) modifications, which determine targeting, stracture, function, and turnover of proteins (Ch. 19). 

Schumacher MC, Resenberger U, Seidel RP, Becker CF, Winklhofer KF, Oesterhelt D, Tatzelt J, Engelhard M (2010) Synthesis of a GPI anchor module suitable for protein post-translational modification. Biopolymers 94 457 -64... 

Wold, F. In Vtuo Chemical Modification of Proteins (Post-Translational Modification). Ann. Rev. Biochem. 50, 788-814 (1981). [A review article on the modified amino acids found in proteins.]... 

Wold, F., In vivo chemical modification of proteins (post-translational modification), Anna. Rev. Biochem., 50,783-814, 1981. 

The mammalian cell lines most commonly employed for prodnction of recombinant proteins are the Chinese hamster ovary and baby hamster kidney cells. These cell lines are nsed in both anchorage-dependent and suspension forms for the prodnction of proteins whose glycoforms are very similar to the native human forms of the corresponding proteins (46). Protein post-translational modifications increase the fnnctional diversity of the... 

A biological membranes system is typically formed by the combination of lipids and proteins. In eukaryotic cells, the plasma membrane, also referred to as the cell membrane, is a protective barrier which regulates what enters and leaves the cell. The endomembrane system is composed of different kinds of membranes which divide the cell into structural and functional compartments within a eukaryotic cell, such as the endoplasmic reticulum, Golgi apparatus, mitochondria, endosome and lysosome. Covalent modification of proteins with lipid anchors (protein lipidation) facilitates association of the lipidated proteins with particular membranes in eukaryotic cells. Protein lipidation is one of the most important protein post-translational modifications (PTMs). Studying lipidated protein function in vitro or in vivo is of vital importance in biological research. 

Post-translational modification, which extends the range of functions of the protein, is one of the later steps in the biosynthesis of many proteins. Post-translational modification of proteins occurs by methylation, hydroxylation, acetylation and phosphorylation of the protein functional groups, by attaching various lipids and carbohydrates to the protein molecule and by making structural changes such as the formation of disulfide bonds from cysteine residues by oxidation. 

Especially promising is the capability of ECD for determining the sites of protein post-translational modifications. Glycosylation and phosphorylation moieties are often labile under standard activation modes. The resulting MS/MS experiments yield little information on modification sites. ECD studies suggest that bonds remote to the modification sites are more labile, and the modification remains intact for MS/MS experiments. This is especially true for modifications such as phosphorylation, ° acetylation,deamidation, ... 

Yukl, E.T. and C.M. Wilmot, Cofactor biosynthesis through protein post-translational modification. Curr. Opin. Chem. Biol, 2012.16(1-2) 54-59. 

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