Proteomics chemical

Chemical proteomics is a parallel investigation of the interactions between small molecule libraries and their binding proteins at the proteome level. 

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Methods based on liquid chromatography-mass spectrometry (LC-MS) and universally accepted search algorithms permit reliable identifications of low levels of proteins at high sensitivity [6]. Even semispecialized protein chemistry labs can readily identify proteins at the level of a few picomoles (10 pmol of a 50-kDa protein is 500 ng). Specialized groups with access to the latest advances in HPLC and mass spectrometry routinely work with subpicomolar quantities. Chemical proteomics as discussed here requires the more advanced equipment. 

The approach recruited to chemical proteomics in Reference [17] is called SILAC (stable isotope labeling with amino acids in cell culture) and is important in comparative proteomics (Figure 1). SILAC works well with cultured mammalian cells, but prokaryotes defeat it by metabolizing the label (usually supplied in lysine and arginine) into other amino acids. For applications beyond cultured eukaryotic cells, the reductive methylation route to differential labeling [18] is among the alternatives [15]-... 

An important factor in all these experiments is the choice of bead used to immobilize the probe. Biochemists have considered cross-linked agarose beads to be exceptionally hydrophilic with a low tendency to bind proteins nonspecifically, and these beads have the further attraction of being commercially available in activated forms (succinimidyl esters, epoxides, and maleimides, for example). However, early trials of bead-based chemical proteomics have shown that many proteins in mammalian cell lysates bind tenaciously to agarose beads. This was unimportant in many studies in which protein-protein interactions were detected by coimmunoprecipitation with immunochemical... 

Activity-based protein profiling (ABPP) is a chemical proteomic strategy in which active-site-directed covalent probes are used to profile the functional states of enzymes in complex proteomes. Activity-based probes (ABPs) can distinguish active enzymes from their inactive zymogens or inhibitor-bound forms. They contain a reactive group intended to modify enzyme active sites covalently and a reporter group (typically rhodamine or biotin) that assists in detection and identification of protein targets. 

Piggott AM, Karuso P. (2004) Quality, not quantity The role of natural products and chemical proteomics in modem dmg discovery. Comb Chem High Throughput Screen 7 607-630. 

Piggott AM, Karuso P. (2008) Rapid identification of a protein binding partner for the marine natural product kahalalide F by using reverse chemical proteomics. ChemBioChem 9 524-530. 

Jeffery DA, Bogyo M. Chemical proteomics and its application to drug discovery. Curr Opin Biotechnol 2003 14 87-95. 

Sem DS, Bertolaet B, Baker B, etal. (2004) Systems-based design of bi-hgand inhibitors of oxidoreductases Filhng the chemical proteomic toolbox. Chem. Biol. 11 185-194. 

Speers AE, Cravatt BF (2005) A tandem orthogonal proteolysis strategy for high-content chemical proteomics. J Am Chem Soc 127 10018-10019... 

Wright AT, Cravatt BF (2007) Chemical proteomic probes for profiling cytochrome p450 activities and drug interactions in vivo. Chem Biol 14 1043-1051... 

Of the approaches listed above the last two (chemical proteomics and ABPP) are currently receiving the most interest and have complementary fields of applications. The affinity-based approach is perfectly suitable for reversible inhibitors, but is limited to quite strong binders. The focus of this chapter will be on the other of these two approaches, namely the activity-based approach to natural product identification. An essential requirement for this approach is that the natural product of investigation contains a reactive functional group that reacts with the protein target, forming a covalent bond. Fortunately, a considerable number of natural products contain such reactive groups [19]. Mostly, electrophilic moieties such as epoxides, Michael acceptors, disulfides, lactones, (3-lactams, quinones, etc. can be found. 

Rix U, Superti-Furga G (2009) Target profiling of small molecules by chemical proteomics. Nat Chem Biol 5 616-624... 

Staub I, Sieber SA (2009) P-Lactam probes as selective chemical-proteomic tools for the identification and functional characterization of resistance associated enzymes in MRSA. J Am Chem Soc 131 6271-6276... 

Heal WP, Wickramasinghe SR, Tate EW (2008) Activity based chemical proteomics profiling proteases as drug targets. Curr Drug Discov Technol 5 200... 

Visser and coworkers combined LC, SPR, and MS together and immobilized cGMP molecules to an SPR chip to monitor the binding and dissociation of proteins from a human lysate by sequential elution steps and SPR [51]. The eluted proteins were thereafter identified by LC-MS/MS. The data indicate that SPR-based chemical proteomics is a viable alternative for quantitative extraction and identification of small-molecule-binding proteins from complex matrices. In fact, SPR-MS technological innovations have grown considerably in the last few years and have even sparked enough interest that leads to a book chapter devoted to the topic [52]. 

Visser NFC, Scholten A, van den Heuvel RHH, Heck AJR (2007) Surface-plasmon-resonan-ce-based chemical proteomics efficient specific extraction and semiquantitative identification of cyclic nucleotide-binding proteins from cellular lysates by using a combination of surface plasmon resonance, sequential elution and liquid chrranatography-tandem mass spectrometry. Chem Bio Chem 7 298-305... 

Daub, H., Godl, K., Brehmer, D., Klebl, B., Muller, G. (2004). Evaluation of kinase inhibitor selectivity by chemical proteomics. Assay Drug Dev. Technol. 2, 215-224. 

Yao H, Costache AD, Sem DS. Chemical proteomic tool for hgand mapping of CYP antitargets An NMR-compatible 3D QSAR descriptor in the heme-based coordinate system. I Chem Inf Comput Sci 2004 44 1456-65. 

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