B-chain of insulin

Neutral endopeptidase (NEP, nephrilysin) is an enzyme that preferentially catalyzes cleavage at the amino group of hydrophobic residues of the B-chain of insulin... 

This lysosomal endopeptidase [EC 3.4.23.5] is similar to pepsin A, except that the specificity is narrower and will not hydrolyze the Gln" —His peptide bond in the B chain of insulin. The enzyme is a member of the peptidase family Al. 

Steps involved in the sequence determination of the B chain of insulin. Amino acids are represented here by their single-letter codes (see table 3.1). 

Fig. 27. Space-filling model of the B chain of insulin. Numbers indicate position of carboxyl oxygens (after Warner231)). The MM in nacreous layer of molluscs is assumed to be built according to the same principles...

In biopolymers hydrates may enforce special conformations141. For example, the harmone oxy tocine of the hypophysis conforms with all hydrophobic groups on one side1271 (Fig. 26) and the hydrophilic groups on the other12 (Fig. 27). Similarity the A-chain and the B-chain of insulin can be arranged to form a disc with a... 

A. M. Blow. Action Of human lysosomal elastase on the oxidized B chain of insulin. Buchan. / 7 7 13(1977). 

The procedure for insulin is more complex because insulin is synthesized as preproinsulin and must be processed to yield the A and B chains of insulin (page 415). Bacteria do not have the processing system for converting the precursor to insulin. The appropriate cDNAs for the A and B chains were coupled into individual plasmids and each inserted into separate bacteria for production of the A and B chains. Following purification of the individual chains/ the proper disulfide bonds were chemically formed to yield the mature insulin. 

Fig. 6. Structura, resemblances between the A and B chain of insulin (beef).

Table VI. Cleavage Sites on S-Sulfo B-Chain of Insulin by Acidic Proteases""...

Table VII. Penicillopepsin Nature of Amino Acids at Cleavage Points in Glucagon and B-Chain of Insulin ...

In an attempt to find out more about the nature of the secondary binding site in penicillopepsin. Mains et al. (76) analysed the nature of the amino acid side chains at positions removed from the sensitive peptide bond. An abbreviated summary of the results is given in Table VII. The number of hydrophobic and hydrophilic side chains respectively are listed for four amino acids on either side of every peptide bond broken in the B-chain of insulin and in glucagon. The positions are numbered Pi, P2, P/, P2 etc. as defined by Berger and Schechter (103). The choice of four positions was taken from the Frutons work (73) on the specificity of pepsin and the eflFect of chain length on catalytic efficiency. The largest eflFects were observed with substrates having three to four... 

This ambiguity was resolved in the experiment shown in Table XL The peptide in this case represents the C-terminal sequence of the B-chain of insulin. The major products were the penta-peptide lacking the N-terminal phenylalanine and free phenylalanine indicating that hydrolysis was the major reaction. In addition, however, there was a significant amount of Phe-Phe. This dipeptide could only come from a transpeptidation involving an acyl transfer. The acyl transfer presumably proceeds via a covalent acyl intermediate. The evidence for this comes from the experiment shown in Table XII where Leu-Tyr-Leu was incubated with both porcine pepsin and penicillopepsin in the presence of a lO-fold excess of C-leucine over Leu-Tyr-Leu. The products, leucine and leucylleucine, were separated by high voltage electrophoresis and analyzed for their specific radioactivity. At most, only traces of radio-... 

Insulin peptides. X. Synthesis of the B-chain of insulin and its combination with natural or synthetic A-chain to generate insulin activity. J. Am. Chem. Soc. 1964 86 930-932. 113. 

To modify the S-carboxymethylated B chain of insulin. Signor et al. (1971) added about 3.0 mmoles of the peptide dissolved in 1.0 ml of 1 M NaOH to 1 ml of water containing 10 mg of nitromalondialdehyde (70 mmoles). After 2 hr at 0-5°C excess modification reagent was removed by gel filtration using 50% formic acid as eluent. 

Human insulin, a hormone that controls blood sugar levels, is made up of two protein chains. Each of the circles represents an amino acid in the protein chains. Disulfide bonds hold the A and B chains of insulin together. Disulfide bonds also join sections of the A chain together. 

The action of chymosin on the B-chain of insulin indicates that it is specific for hydrophobic and aromatic amino acid residues (Fish, 1957). Chymosin is relatively weakly proteolytic indeed, limited proteolysis is one of the characteristics to be considered when selecting proteinases for use as rennet substitutes (Fox, 1989a). 

Insulin interacts with its cell surface receptor via key amino acid residues located along the N- and C-termini of the A chain of insulin and along the carboxy terminus of the B chain of insulin (Table 32.6). The binding of insulin occurs to amino acid residues located within the N- and C-terminal regions of the a subunit of the receptor, which includes a cysteine-rich region (5). Binding and activation of the insulin receptor results in a cascade of biochemical events previously described. 

The neutral mutation rate differs for each nucleotide in a gene and usually serves as a good indicator for the functional importance of encoded amino acid residue. For example, the C-peptide of proinsulin evolves at a rate (0.526PAM/10 years) much more rapid than that of the A- and B-chains of insulin (0.071 PAM(point accepted mutation)/10 years) because the C-peptide appears to promote the protein folding and is removed pro-teolytically after the insulin has folded to its correct conformation. The greater divergent rate of the C-peptide than that of the A and B-chains reflects the fewer constraints on its... 

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