Disulphide bond, reduction

Table IV. Methods Provided for Disulphide Bond Reduction and Alkylation in ABRF-94SEQ...

A development reported recently [519] involves reduction of the cystine disulphide bonds in wool with either thioglycolic acid or tetrakis(hydroxymethyl)phosphonium chloride to form thiol groups, followed by crosslinking with bifunctional reactive dyes. This gave improved insect resistance but had adverse effects on physical properties such as strength, shrinkage and stiffness, thus limiting the potential of the process for commercial use. 

The method described here is based on two reactions first, the reduction of the disulphide bond between Hey and other thiols or the cysteine residue of proteins by the reducing compound tri-n-butylphosphine (Fig. 2.2.2a) followed by the reaction of Hey and other thiols with the flourogenic thiol-specific reagent ammonium... 

Reductants provide a means to fully denature proteins within the sample and aid solubilization. To fully break disulphide bonds,... 

Polarography has been successfully applied to the investigation of structural problems involving sulphur compounds. The presence of a disulphide bond has been established by means of the polarographic reduction waves of cytochrome C (156) and lipoic acid (757), and in cyclic disulphides of the oxytocine and vasopressine type (158). The elucidation of the process responsible for the reduction wave of lipoic acid was carried out by comparison with reduction waves of cyclic disulphides, where the disulphide bond was incorporated into rings of various size. The similarity indicated that in lipoic acid an S—S bond which is a part of a larger cyclic system is reduced. 

Bockle, B. and Muller, R. (1997). Reduction of disulphide bonds by Streptomyces pactum during growth on chicken feathers. Appl. Environ. Microbiol. 63,... 

Another possibility to introduce two functional groups at a predetermined distance was investigated by polymerising the disulphide 6 under imprinting conditions (see entry k). Subsequent reduction of the disulphide bond with diborane provides polymers with two closely positioned mercapto groups (see Scheme 4.I1I). 

Free formaldehyde is released from the above reaction, which in turn acts as a reducing agent on the polymer disulphide bonds to bring about a reduction into thiol with the formation of formic acid ... 

Out of the above bonds, the disulphide bond (covalent bond) is the strongest and cannot be affected by solvent, pH, temperature and salts whereas the above conditions. The disulphide bond can be split and reformed by oxidation/reduction respectively. The tertiary structure gains special importance in the case of enzymes. 

It is usual to effect cleavage of disulphide bonds by reduction or oxidation. Addition of a large excess of a thiol such as 2-mercaptoethanol or 1,4-dithiothreitol to a polypeptide reduces cystine residues to cysteine (Scheme 5.1). In order to prevent reoxidation in air, the generated thiol groups are blocked, usually by reaction with iodoacetic acid. The product yields S -carboxymethylcysteine (5.9) on hydrolysis for amino-acid analysis. Alternatively, oxidative cleavage of disulphide bonds can be achieved with performic acid each half of the cysteine residue is converted into a residue of cysteic acid (5.10). 

Finally, disulphide bonds can be located by hydrolysing a protein to a mixture of peptides using either a proteinase or a specific chemical method of cleavage and the mixture can be analysed directly by fast-atom bombardment mass spectrometry (Chapter 3) and again after reduction of disulphide bonds (Yazdanparast et al., 1987). By identifying those peaks which disappear as a result of reduction and new peaks with appropriate masses that have taken their place, it is simple to assign disulphide bonds to the relevant amino-acid sequences. 

Other chemical degradation pathways include -elimination, which can lead to racemization and disulphide exchange reactions. The amino acids that may undergo P-elimination include Cys, Ser, Thr, Phe, and Lys (40) and occur especially at alkaline pH (64). The reduction and oxidation of the disulphide bonds are often accompanied by a considerable change in the protein conformation (23,33). An example is the secondary structure of insulin that is disrupted or completely lost when the disulphide bonds are broken (23). The disulphide bond disruption or interchange can also result in an altered three-dimensional structure and therefore a possible loss of activity (40) or aggregation (29). For further details, the reader is directed to the following reviews and book chapters (30,40,62,65). 

An interesting aspect is that moths only thrive on keratin which contains disulphide bonds. If these are ruptured by reduction with sodium hydrosulphite and thioglycoUic acid and new links are built by methods such as joining primary amino groups with glyoxal, the wool is immune from attack by moths. 

P. A. Price, W. H. Stein, and S. Moore. Effect of divalent cations on the reduction and reformation of the disulphide bonds of deoxyribonuclease. J. Biol. Chem. 244 929-932 (1969). 

For preparing whole ricin IT, the intact toxin is normally derivatized with a bifunctional cross-linking agent before being mixed with antibody prepared for cross-linking by partial reduction of disulphide bonds [183]. 

A putative y-amide derivative was also prepared from MTX by reaction with cystamine in water solution in the presence of EDC hydrochloride, followed by reduction of the disulphide bond with excess 2-mercaptoethanol [328]. The resulting thiol (VIII.220) was then allowed to react with... 

Other time-resolved synchrotron radiation scattering studies have been performed to study the dissociation of aspartate transcarbamylase by mercurials. These showed that a time resolution of the order of 100 msec is possible [142aj. The aggregation of bovine serum albumin after cleavage of its disulphide bonds by reduction with dithiothreitol could be traced in time intervals of 50 sec and also showed that experiments with a time interval of 100 msec are possible [142b],... 

Oxidation of EtsP proceeds by reduction of albumin disulphide bonds (SSA = any albumin species) see equation 60. 

Sheep erythrocytes sensitized with intact immunoglobulin G or with reduced (dithioerythritol) and alkylated (iodoacetamide) antibody have been lysed by guinea-pig serum, indicating that reduced and alkylated antibody bound and activated complement. Observations suggested that the sole effect of reduction of antibody disulphide bonds was to diminish the co-operativity of antibody-complement interaction. 

The effects of dimethyl sulphoxide, lithium bromide, guanidinium chloride, sodium dodecyl sulphate, and urea on lysozyme have been studied using Raman spectroscopy. The spectrum observed was found to depend on the denaturant used, suggesting there is not a unique denatured state for lysozyme. An analysis of the interaction of sodium dodecyl sulphate with lysozyme has been published. A kinetic study of the denaturation and subsequent reduction of disulphide bonds in lysozyme has been made using rapid ultrasonic absorption measurements. 

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