Proteins enzyme cleavage

Amyloid precursor protein (APP) is the precursor of (3-amyloid, the main component of senile plaques found in the brain of Alzheimer patients. The production of (3-amyloid from APP to the cells from abnormal proteolytic cleavage of the amyloid precursor protein. Enzymes involved in this cleavage may be suitable targets for the therapy of Alzheimer s disease. 

It is clear that both intact cell MALDI-TOF and PFGE have their limitations. PFGE analyses probes the chromosomal DNA of microorganisms for variations in the locations of specific restriction enzyme cleavage sites, while MALDI-TOF mass spectrometry of intact cells primarily examines abundant proteins such as ribosomal proteins35 and those associated with or near bacterial cell walls.58 In order for MALDI-TOF to detect a variation, a mutation must lead to noticeable changes in the expression of cell wall—associated... 

A third approach is to completely synthesize a gene for the protein under study. The sequence does not have to be exactly the natural one but can be made with restriction enzyme cleavage sites that permit easy excision and readdition of particular fragments. Synthetic fragments containing various mutations can then be grafted in at will. Genes of this type have been made for rhodopsin and related proteins (Chapter 23) and for numerous other proteins. 

Tryptic peptide mapping. The technique of generating a chromatographic profile characteristic of the fragments resulting from trypsin enzyme cleavage of the protein. 

Somatomedins are synthesized as larger precursors, the structures of which in man have been determined by cloning corresponding cDNAs and nucleotide sequence determination [53,54]. The precursors are processed by enzymic cleavage to give the mature somatomedins, which are secreted from the cells of origin. There is little storage of somatomedins in the cells where they are produced they are probably secreted from the cells in vesicles by the same route followed by other secreted hepatic proteins such as serum albumin. 

Based on chemical considerations alone, ribozymes should be able to catalyze many different types of reactions. Ribozymes can maintain defined secondary and tertiary structures, just as protein enzymes do. Ribozymes can interact with substrates specifically via hydrogen bond networks, just as protein enzymes do. Finally, ribozymes have available to them a chemistry that, while more limited than of proteins, is substantial. RNA contains proton donors and acceptors with pK, values that cluster at 4 and 9.71 The critical lack of a good donor/acceptor with a pKt near 7 can be rectified by any of several simple expedients, such as modification of guanosine to 7-methylguanosine, protonation of triple base-paired cysto-sine,72 or inclusion of a proton donor/acceptor in an environment with a different polarity than water (in this respect, it is interesting to note that Dahm and Uhlenbeck have found that the cleavage reaction catalyzed by the hammerhead ribozyme is dependent on some dissociable proton with a pKa of 8.0).73... 

If your protein sample has an acetylated N-terminus, it will not yield any information from Edman degradation of the intact protein. Enzymes have been described that remove N-acetyl groups from peptide chains, but, in general, these have been found to function well only on peptides and not on intact proteins. Procedures have been described for cleavage of blocked proteins into peptides followed by modihcation of all newly revealed peptide N-termini (e.g., by succiny-lation) and then incubation of the mixture with a deacetylase. The entire mixture can then be subjected to Edman degradation. Only the deacetylated peptide representing the original N-terminus will provide sequence. 

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