Protein-small molecule interactions/ complexes

Proteins, not mRNAs, are the true functional components of cells. Unlike DNA microarrays, on which interactions are based on Watson-Crick base pairing, biomo-lecular interactions on protein microarrays are determined by complex associations between the probe proteins and the target molecules. Individual protein-ligand pairs could differ greatly in their affinities. Furthermore, unlike DNA whose structure is relatively simple, proteins are extremely diverse in structure and functions, and often display many variables, such as posttranslational modifications. Protein microarrays are useful for determining numerous protein interactions including protein-protein [59], protein-DNA [26], and protein-small molecule interactions [30], or identifying the substrates of protein kinases [58]. 

Inspired by these innovative applications of fluorous tags, we have developed a fluorous PAL technique to probe protein-small molecule interactions. In this approach (Fig. 1), a fluorous tag is incorporated into the PAL reagents to allow specific enrichment of labeled components fi om the complex assay mixture [23-25] for MS analysis. In a typical PAL experiment, as illustrated in Fig. I,... 

The general types of protein-protein interactions that occur in cells include receptor-ligand, enzyme-substrate, multimeric complex formations, structural scaffolds, and chaperones. However, proteins interact with more targets than just other proteins. Protein interactions can include protein-protein or protein-peptide, protein-DNA/RNA or protein-nucleic acid, protein-glycan or protein-carbohydrate, protein-lipid or protein-membrane, and protein-small molecule or protein-ligand. It is likely that every molecule within a cell has some kind of specific interaction with a protein. 

Molecular Interactions. Various polysaccharides readily associate with other substances, including bile acids and cholesterol, proteins, small organic molecules, inorganic salts, and ions. Anionic polysaccharides form salts and chelate complexes with cations some neutral polysaccharides form complexes with inorganic salts and some interactions are stmcture specific. Starch amylose and the linear branches of amylopectin form inclusion complexes with several classes of polar molecules, including fatty acids, glycerides, alcohols, esters, ketones, and iodine/iodide. The absorbed molecule occupies the cavity of the amylose helix, which has the capacity to expand somewhat to accommodate larger molecules. The starch—Hpid complex is important in food systems. Whether similar inclusion complexes can form with any of the dietary fiber components is not known. 

Separation media, with bimodal chemistry, are generally designed for the complete separation of complex samples, such as blood plasma serum, that typically contain molecules differing in properties such as size, charge, and polarity. The major principle of bifunctional separation relies on the pore size and functional difference in the media. For example, a polymer bead with hydrophilic large pores and hydrophobic small pores will not interact with and retain large molecules such as proteins, but will interact with and retain small molecules such as drugs and metabolites. 

The abundance of structural information has led to a significant increase in the use of structure-based methods both to identify and to optimise inhibitors of protein kinases. The focus to date has centred upon small molecule ATP-competitive inhibitors and there are numerous examples of protein-ligand complexes available to guide design strategies. ATP binds in the cleft formed between the N- and C-terminal lobes of the protein kinase, forming several key interactions conserved across the protein kinase family. The adenine moiety lies in a hydrophobic region between the jS-sheet structure of subdomains I and II and residues from subdomains V and VIb. A... 

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