Intramolecular disulphide bridge

It is secreted by acidophil cells. Human growth hormone has a single straight chain polypeptide structure containing two intramolecular disulphide bridges and is composed of 188 amino acids. 

GOase consists of a single polypeptide chain with two intramolecular disulphide bridges (Kosman et al., 1974). It is reported to be fully active in 6M urea (Kosman et al., 1974) attesting to its stability and potential for biotechnological applications. 

D-Xylosidase from Bacillus pumilus has been shown to contain one intramolecular disulphide bridge and four L-cysteinyl residues per monomer (mol. wt. 6.0 X 10 ). Under native conditions, one L-cysteinyl residue per polypeptide chain reacted with 5,5 -dithiobis(2-nitrobenzoic acid) or 4-chloro-mercuribenzoate resulting in complete loss of enzymic activity. Reductive treatment with dithiothreitol completely reversed this loss. Kinetic analysis of the inactivation reaction pointed to a saturation effect, occurring when the concentration of the — SH-directed reagent was increased. A reaction model, which included prior association of the reagent with the enzyme, was proposed. 

Among all possible configurations, the active enzyme conformation is assumed to be thermodynamically the most stable in situ. Experimental evidence to validate this assumption came from reversible denaturation studies on enzymes. For example. ribonuclease may be completely unfolded in urea solution after breakage of four native intramolecular disulphide bridges. Afterwards, when ribonuclease is brought to renature in the proper conditions, the native disulphide bonds are restored, and the protein recovers its catalytic activity. This case must be regarded, however, as a very favourable one because in most instances renaturation is... 

The insulin molecule consists of two chains, the A-chain with 21 amino acids and the B-chain with 30 amino acids (Fig. 12). They are interconnected by two intermolecular disulphide bridges between amino acids A7 and B7 and A20 and B19. A third disulphide bridge connects amino acids 6 and 11 on chain A, giving an intramolecular loop. It is synthesized as a single-chain precursor, preproinsulin, which is converted to proinsulin after the molecule has been translocated to the endoplasmic reticulum. There, the C-peptide, which connects the A- and B-chains, is cut away forming the active insulin (Briggs and Gierasch, 1986 Bailyes etal., 1993). The most often used insulins in therapeutics (Fig. 12), bovine, porcine and human insulin, exhibit differences in their amino acid sequences bovine insulin contains Ala instead of Thr in position 8 and Val instead of lie in position 10 of the A-chain, and both bovine and porcine insulin differ from human insulin by an Ala instead of Thr in position 30 of the B-chain. 

This class of peptides typically contains 18 or 19 amino acid residues and share a common 13 amino acid sequence, which is Cys -Cys-Glu-Leu-Cys-Cys °-Asn-Pro-Ala-Cys-Ala -Gly-Cys (full peptide position) and Cys -Cys-Glu-Leu-Cys-Cys -Asn-Pro-Ala-Cys-Ala -Gly-Cys (toxic domain). Since the domain is conserved in several enterotoxins, one expects this 13 residue domain to be the primary reason for the toxicity. The six Cys residues form three disulphide bridges, and provide stability to the structure of the protein. lETN has a simple structure it has got three beta ((3) turns. In addition, the crystal structure contains five intramolecular hydrogen bonds that also add to the stability of the protein. Another interesting aspect of this protein is the presence of 13 water molecules in the crystal structure (Sato et al., 1986). Overall, this protein has a hydrophobic character, as the side chains of all residues are oriented to the outside of the molecule (Balasubramanian et al., 2003). 

Scheme 1. Synthetic approaches to the formation of disulphide bridges. The residues to be linked are either on the same (intramolecular reactions dotted line indicates intervening residues) chain or different (intermolecular reaction dotted line indicates discontinuity) chains. and Xj designate S-protecting groups that are stable to chain assembly conditions. Xj and Xj may or may not be the same, depending on the chemistry chosen for the directed method. Reactions can be carried out either on-resin or in solution this facet and additional considerations are described more fully in the text.

To gain biological activity the de novo synthesized polypeptide must acquire a specific three-dimensional configuration. The bonds which determine this conformation are of two types firstly, the intramolecular covalent disulphide bridges, and secondly, weak bonds, hydrogen bonds, van der Waals interactions and ionic bonds. 

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