Escherichia coli disulfide bonds

Oudot, C. Jaquinod, M. Gortay, J.C. Cozzone, A.J. Jault, J.M. The isocitrate dehydrogenase kinase/phosphatase from Escherichia coli is highly sensitive to in-vitro oxidative conditions role of cysteine67 and cysteinel08 in the formation of a disulfide-bonded homodimer. Eur. J. Biochem., 262, 224-229 (1999)... 

A major class of insoluble proteins are recombinant proteins expressed (usually in Escherichia coli) as inclusion bodies. These are dense aggregates found inside cells that consist mainly of a desired recombinant product, but in a nonnative state. Inclusion bodies may form for a variety of reasons, such as insolubility of the product at the concentrations being produced, inability to fold correctly in the bacterial environment, or inability to form correct, or any, disulfide bonds in the reducing intracellular environment. Their purification is simple, since the inclusion bodies can be separated by differential centrifugation from other cellular constituents, giving almost pure product the problem is that the protein is not in a native state, and is insoluble. Some methods for obtaining an active product from inclusion bodies are described in Coligan et al. (2001). 

Bessette, P. H., F. Aslund, J. Beckwith and G. Georgiou. Efficient folding of proteins with multiple disulfide bonds in the Escherichia coli cytoplasm. Proc Natl Acad Sd U S A (1999) 96(24) 13703-8. 

Fuh, G., Mulkerrin, M. G., Bass, S., McFarland, N., Brochier, M., Bourell, J. H., Light, D. R., and Wells, J. A. (1990). The human growth hormone receptor. Secretion from Escherichia coli and disulfide bonding pattern of the extracellular binding domain./ Biol. Ghent. 265(6), 3111-3115. 

Fischer, B. Summer, L Goodenough, P. Isolation, renaturation and formation of disulfide bonds of eukaryotic proteins expressed in Escherichia coli as inclusion bodies. Biotechnol. Bioeng. 1993, 41, 3-13. 

Fig. 31. Reversible inactivation of Fi (p Asp-380->Cys) by formation of a specific p y intersubunit disulfide bond. Wild-type (w.t.) and mutant F, samples were treated with DTNB and aliquots were taken for SDS-PAGE and ATPase assays. See text for other details. Figure source Duncan, Bulygin, Zhou, Hutcheon and Cross (1995) Rotation of subunits during catalysis by Escherichia coli F,-ATPase. Proc Nat Acad Sci, USA 92 10966.

R Aggeier, MA Houghton and RA Capaidi (1995) Disulfide bond formation between the COOH-terminal domain of the p subunits and the y and e subunits of the Escherichia coli F -ATPase. J Bioi Chem 270 185-9191 i Ogiivie, R Aggeier and RA Capaidi (1997) Cross-linking of the S-subunit to one of the three a subunits has no effect on functioning, as expectged ifS is a part of the stator that links the F, and Fq parts of the Escherichia coli ATP synthase. J Bioi Chem 272 16652-16656... 

CATALYSIS OF DISULFIDE BOND FORMATION AND ISOMERIZATION IN Escherichia coli... 

Derman, A. I., Prinz, W. A., Belin, D., and Beckwith, J. (1993). Mutations that allow disulfide bond formation in the cytoplasm of Escherichia coli. Science 262, 1744-1747. 

Di Abietes. Di Abietes treatment was first changed to daily injections j of Humulin instead of beef insulin. Humulin is now mass-produced by recombinant DNA techniques that insert the human DNA sequences for the insulin A and B chains into the Escherichia coli or yeast genome (see Chapter 17). The insulin chains that are produced are then extracted from the media and treated to form the appropriate disulfide bonds between the chains. As costs have fallen for production of the synthetic human insulins, they have replaced pork insulin and the highly antigenic beef insulin. 

Ostermeier M, De Sutter K, Georgiou G. Eukaryotic protein disulfide isomerase complements Escherichia coli dsbA mutants and increases the yield of a heterologous secreted protein with disulfide bonds. J Biol Chem 1996 27l(18) 106l6-10622. 

Missiakas, D., Georgopoulos, C., and Raina, S. (1994) The Escherichia coli dsbC (xprA) gene encodes a periplas-mic protein involved in disulfide bond formation. EMBO J., 13, 2013-2020. 

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