Proteins enzymatic fragmentation

Brash has also used CD to study eluted proteins and finds large changes in a-helix content of fibrinogen, perhaps due to enzymatic fragmentation produced by the surface-induced activation of plasminogen to plasmin11 18). 

A considerable amount of information is available on the reversible, pH-dependent molecular conformational alterations which occur in both bovine and human serum albumins. This work has yielded valuable knowledge regarding the structural organization of the albumin molecule and has been instrumental in devising conditions and procedures for the enzymatic fragmentation of the protein and is therefore pertinent to an immunochemical consideration. 

Coagulation Factors II, III, VII, IX, X, XI, and Xlla fragments, thrombin, and plasmin are classified as serine proteases because each possesses a serine residue with neighboring histidine and asparagine residues at its enzymatically active site (Table 3). Factors II, VII, IX, and X, Protein C, Protein S, and Protein Z are dependent on the presence of vitamin K [84-80-0] for their formation as biologically functionally active procoagulant glycoproteins. 

Such a process led to the full assignment of the sequence of the cytochrome c" protein shown in Figure 5.17, in which the polypeptide fragments generated by enzymatic digestion with endoproteinase Lys-C and Glu-C are also indicated. 

Subsequently, proteolytic fragments of the rabbit renal 25-kDa amiloride-binding protein were micro-sequenced and found to have high sequence homology with rat and human NAD(P)H quinone oxidoreductase. Indeed, enzymatic assays revealed that renal brush border membrane vesicles contain significant NADPH quinone oxidoreductase activity. Presumably NAD(P)H quinone oxidoreductase coincidentally binds amiloride analogs with the same rank order as the Na /H exchanger [39]. 

Another method that has been applied by our group to the study of enzymatic reactions is the Effective Fragment Potential (EFP) method [19]. The EFP method (developed at Mark Gordon s group at Iowa State University) allows the explicit inclusion of environment effects in quantum chemical calculations. The solvent, which may consist of discrete solvent molecules, protein fragments or other material, is treated explicitly using a model potential that incorporates electrostatics, polarization, and exchange repulsion effects. The solute, which can include some... 

A purified protein having the amino acid sequence for ORF-1 (SEQ ID No 2) or an enzymatically active fragment thereof. 

The fragments formed by FeBABE fragmentation are analyzed by comparing them to enzymatic or chemical cleavage patterns observed by treatment on the same prey protein. Since the cleaved prey protein is detected by its end-labeled tag, the only fragments detected are those... 

Analyze the cleaved proteins by SDS gel electrophoresis and Western blotting, followed by specific detection of the prey protein label. Comparison of prey protein fragments formed by chemical and enzymatic digestion can yield information as to the site of the resultant FeBABE-mediated cleavage. 

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