Cleavage of disulfides

FIGURE 5.18 Methods for cleavage of disulfide bonds in proteins, (a) Oxidative cleavage by reaction with performic acid, (b) Reductive cleavage with snlfliydryl compounds. Disulfide bridges can be broken by reduction of the S—S link with snlfliydryl agents such as 2-mercaptoethanol or dithiothreitol. Because reaction between the newly reduced —SH groups to re-establish disulfide bonds is a likelihood, S—S reduction must be followed by —SH modification (1) alkylation with iodoac-etate (ICH,COOH) or (2) modification with 3-bromopropylamine (Br— (CH,)3—NH,). 

Cleavage of disulfide bonds is done easily by incubation with a reducing agent at a level of 10-100 mM concentration. If the disulfides in the crosslinks are the only ones present in the complexed molecules, then reduction will yield unconjugated molecules—one or both of... 

Figure 8.1 Cleavage of disulfide-containing crosslinking compounds can be done using a reducing agent such as DTT. Reduction causes the conjugates to break apart into their original components with each component containing a portion of the crosslinker that terminates in a thiol group.

Chemical properties of disulfide dications are almost similar to the properties of disulfonium dications. Nucleophilic exchange is characteristic transformation of disulfide dications. Substitution occurs at sulfur atom with cleavage of disulfide moieties because of the lower energy of S-S bonds (Scheme 59). 

Example The cleavage of disulfide bonds by reduction with 1,4-dithiothreitol causes the unfolding of the protein. This exposes additional basic sites to protonation, and therefore results in higher average charge states in the corresponding ESI spectrum (Fig. 11.14). [88]... 

Dai S, Schwendtmayer C, Schiirmann P, et al. 2000. Redox signaling in chloroplasts cleavage of disulfides by an iron-sulfur cluster. Science 287 655-8. 

Fig.1 S - S-bond cleavage of disulfides on gold (a) and tilting of thiol chains on gold (b)...

Reductive cleavage of disulfides is very easy and can be accomplished without affecting some other readily reducible groups such as nitro groups. 

Polymer supported persulfonic acids can be prepared by treating polymer-bound sulfonic acids with H2O2 or K2S20g. The resulting resin was found to display an activity of 2.5 mole equivalents per gram of wet resin. This persulfonated resin was successively applied for the oxidation of carboxylic acids, ketones, olefins and for the cleavage of disulfide linkage and of A-formylamino acids. 

Polymer-supported persulfonic acid was prepared from potassium persulfate and the cation exchange resin P—SO3H in water. The authors reported various applications of this new oxidizing reagent such as epoxidation of olefins, Baeyer-Villiger reaction and cleavage of disulfide and Af-formylamino acids. 

Mix samples with one-third of their volume of buffer G and heat at about 70 °C for 5 min. For cleavage of disulfide bridges add DTT to a final concentration of 10 mM. 

Cleavage of Disulfide Bridges 147 5. Other Information from NMR Spectra... 

It has been generally accepted that the extreme-pressure performance of disulfides is better than that of monosulfides (Allum Ford 1965 Forbes 1970). The difference was explained simply with the anion radical lubrication model (Kajdas 1994). Monosulfides are reduced less readily than disulfides. Reductive cleavage of disulfides with the generation of active species RS and RS proceeds more easily than that of monosulfides. Accordingly, disulfides exhibit more efficient load-carrying properties. 

Pardridge, W.M., D. Triguero, and J.L. Buciak. 1990. Beta-endorphin chimeric peptides Transport through the blood-brain barrier in vivo and cleavage of disulfide linkage by brain. Endocrinology 126 977. 

The anodic cleavage of disulfides was used on the laboratory scale for the synthesis of other vulcanization enhancers 330 331), for the production of phenyl sulfinates 332), and for the synthesis of intermediates for penicillins and cephalosporins 333). 

Cleavage of disulfides.1 Diaryl disulfides are cleaved to thiols by semicarbazide in CH2C12-CH30H at room temperature within 2 hours. Cleavage of di-heteroaryl disulfides proceeds more readily than that of diaryl disulfides. 

Disulfide linkages may be broken either oxidatively or reductively. The former method involves the treatment of the protein with performic add, which converts all disulfide bonds into cysteic add residues. This procedure is usually performed before a protein is hydrolyzed for amino add analysis. Cystine and cysteine are then determined as cysteic add. The reductive cleavage of disulfide bonds involves the treatment of the protein with mercaptoethanol (SH-CH2-CH2-OH), followed by the alkylation of the newly formed -SH groups. The complete disruption of all secondary interactions (that is, complete denaturation) can be achieved in most proteins with 6 M guanidine hydrochloride and 0.1 M mercaptoethanol or 8 M urea and 0.1 M mercaptoethanol. 

Unlike alcohols, thiols are easily oxidized to give a dimer called a disulfide. The reverse reaction, reduction of the disulfide to the thiol, takes place under reducing conditions. Formation and cleavage of disulfide linkages is an important aspect of protein chemistry (Chapter 24), where disulfide bridges between cysteine amino acid residues hold the protein chain in its active conformation. More examples of disulfide bridges appear in Section 24-8C. 

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