Protein binding reactions

T. M. Li and J. F. Burd, Enzymic hydrolysis of intramolecular complexes for monitoring theophylline in homogeneous competitive protein-binding reactions, Biochem. Biophys. Res. Commun. 103, 1157-1165 (1981). 

Baker, B.M. and K.P. Murphy. 1996. Evaluation of linked protonation effects in protein binding reactions using isothermal titration calorimetry. Biophys J 71 2049-2055. 

Electronic complexity reduction may provide an alternative method for sequence enrichment that is rapid, user-friendly and potentially quantitative. The device used in this experiment permits very high current densities and thus allows transport in buffers other than those typically used for electrophoresis. Beyond the use in complexity reduction, this device, with its ability to sustain high current densities, may have application in hybridization assays with a limited number of probes, immunoassays or other protein-binding reactions, and cell transport studies. Furthermore, the use of electrophoretic transport through all of the steps from sample processing through the assay should facilitate systems integration. 

Protonation Effects in Protein Binding Reactions Using Isothermal Titration Calorimetry. 

The determination of the quantity of protein bound to the insoluble carrier sometimes causes difficulties. The methods usually applied are laborious or somewhat inaccurate. Labeling of assayed protein, for instance with C-acet-anhydride, makes it possible to carry out a very fast and exact determination of immobilized protein The determination of bound enzyme C-labeled aldolase after its immobilization on polyacrylamide can serve as an example The concentration measurements of certain proteins are based on their ability to bind certain ligands. Radiolabels such as or H-biotin have been used for the determination of avidin by direct binding or for biotin assay by isotopic dilution Cofactor and fluorescent labeled ligands have been also used for the monitoring of specific protein binding reactions. 

Viadiu H, Aggarwal AK. Structure of BamHI bound to nonspecific DNA a model for DNA sliding. Molec. Cell 2000 5 889-895. Sidorova NY, Muradymov S, Rau DC. Trapping DNA-protein binding reactions with neutral osmolytes for the analysis by gel mobility shift and self-cleavage assays. Nucleic Acids Res. 2005 33 5145-5155. 

Burd, J.F., Carrico, R.J., Fetter, M.C., Buckler, R.T., Johnson, R.D., Bo-guslaski, R.C. and Christner, J.E. (1977). Specific protein-binding reactions monitored by enzymatic hydrolysis of ligand-fluorescent dye conjugates. Anal. Biochem. 77, 56-57. 

Metallointercalators for DNA and Polymeric Molecular Switch for Protein Binding Reactions... 

Sulfaphenazole (684) and sulfazamet (685) are both examples of relatively short acting sulfonamides (B-80MI40406) and their antibacterial activity has been tested against Escherichia coli, the former being more effective than the latter. Sulfaphenazole also displaces sulfonyl ureas from protein binding sites on human serum albumin and consequently increases the concentration of the free (active) drug and produces a more intense reaction that may result in hypoglycemia. 

One target type for which the molecular mechanism of efficacy has been partly elucidated is the G-protein-coupled receptor (GPCR). It is known that activation of GPCRs leads to an interaction of the receptor with separate membrane G-proteins to cause dissociation of the G-protein subunits and subsequent activation of effectors (see Chapter 2). For the purposes of binding, this process can lead to an aberration in the binding reaction as perceived in experimental binding studies. Specifically, the activation of the receptor with subsequent binding of that... 

PTKs can be subdivided into two large families, receptor tyrosine kinases (RTKs) and non-RTKs. The human genome encodes for a total of 90 tyrosine kinases of which 32 are nonreceptor PTKs that can be placed in 10 subfamilies (Fig. 1). All nonreceptor PTKs share a common kinase domain and usually contain several additional domains that mediate interactions with protein-binding partners, membrane lipids, or DNA (Table 1). These interactions may affect cellular localization and the activation status of the kinase or attract substrate proteins for phosphorylation reactions. 

Such free radicals may be stabilized by binding to proteins. Redox reactions may also occur between ionic species, for example the oxidation of reduced cytochrome c by hexacyanoferrate (ferricyanide) ions. 

Finally, the binding of specific transcription factors to cognate DNA elements may result in disruption of nucleosomal structure. Many eukaryotic genes have multiple protein-binding DNA elements. The serial binding of transcription factors to these elements—in a combinatorial fashion—may either directly disrupt the structure of the nucleosome or prevent its re-formation or recruit, via protein-protein interactions, multiprotein coactivator complexes that have the ability to covalently modify or remodel nucleosomes. These reactions result in chromatin-level structural changes that in the end increase DNA accessibifity to other factors and the transcription machinery. 

This equation illustrates the components of a competitive protein binding assay system. That is, the reaction system contains both radioactive and non-radioactive free ligand (P and P) and both radioactive and non-radioactive protein bound ligand (P Q and PQ). This type of assay assumes that binding protein will have the same affinity for the labeled or non-labeled material that is being measured. Although this assumption is not always completely valid, it usually causes no problems of consequence with most radioassays or radioimmunoassays. 

---