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Review
. 2009 Dec;10(6):614-25.
doi: 10.2174/138920309789630534.

The role of thiols and disulfides on protein stability

Maulik V Trivedi  1 , Jennifer S Laurence, Teruna J Siahaan
Affiliations
Review

The role of thiols and disulfides on protein stability

Maulik V Trivedi et al. Curr Protein Pept Sci. 2009 Dec.

Abstract

There has been a tremendous increase in the number of approved drugs derived from recombinant proteins; however, their development as potential drugs has been hampered by their instability that causes difficulty to formulate them as therapeutic agents. It has been shown that the reactivity of thiol and disulfide functional groups could catalyze chemical (i.e., oxidation and beta-elimination reactions) and physical (i.e., aggregation and precipitation) degradations of proteins. Because most proteins contain a free Cys residue or/and a disulfide bond, this review is focused on their roles in the physical and chemical stability of proteins. The effect of introducing a disulfide bond to improve physical stability of proteins and the mechanisms of degradation of disulfide bond were discussed. The qualitative/quantitative methods to determine the presence of thiol in the Cys residue and various methods to derivatize thiol group for improving protein stability were also illustrated.

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Figures

Figure 1
Figure 1
Increasing trend in the use of recombinant proteins and monoclonal antibodies as drugs. The number of new biotech drugs and new indications approved for biotech drugs from 1982 to 2005.
Figure 2
Figure 2
Various labeling reagents for the detection and quantification of protein thiols and disulfides. (a) Labeling protein thiols with 5,5'-dithio-bis(2-nitrobenzoic acid) (DTNB or Ellman’s reagent), 4,4'-dithiodipyridine (DTDP), n-octyl-5-dithio-2-nitrobenzoic acid (ODNB), and ThioGlo®1. (b) Labeling cellular protein thiols with bimanes: monobromobimane (mBBr) and dibromobimane (bBBr) label thiols inside cells as well as on cell surfaces, whereas monobromotrimethylammoniobimane (qBBr) labels only the cell surface protein thiols.
Figure 2
Figure 2
Various labeling reagents for the detection and quantification of protein thiols and disulfides. (a) Labeling protein thiols with 5,5'-dithio-bis(2-nitrobenzoic acid) (DTNB or Ellman’s reagent), 4,4'-dithiodipyridine (DTDP), n-octyl-5-dithio-2-nitrobenzoic acid (ODNB), and ThioGlo®1. (b) Labeling cellular protein thiols with bimanes: monobromobimane (mBBr) and dibromobimane (bBBr) label thiols inside cells as well as on cell surfaces, whereas monobromotrimethylammoniobimane (qBBr) labels only the cell surface protein thiols.
Figure 3
Figure 3
Degradation reactions of protein disulfide bonds in neutral and basic conditions. (a) Direct attack on sulfur atom by hydroxyl anion, (b) β-elimination reaction, and (c) α-elimination reaction.
Figure 3
Figure 3
Degradation reactions of protein disulfide bonds in neutral and basic conditions. (a) Direct attack on sulfur atom by hydroxyl anion, (b) β-elimination reaction, and (c) α-elimination reaction.
Figure 3
Figure 3
Degradation reactions of protein disulfide bonds in neutral and basic conditions. (a) Direct attack on sulfur atom by hydroxyl anion, (b) β-elimination reaction, and (c) α-elimination reaction.
Figure 4
Figure 4
Reagents for derivatization of free thiols in proteins.
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