Disclosed is a method for refolding a protein or peptide that does not contain essential disulfides and that contains at least one free cysteine residue. Also disclosed are polymer IFN- conjugates that have been created by the chemical coupling of polymers such as polyethylene glycol moieties to IFN-, particularly via a free cysteine in the protein. Also disclosed are analogs of bioactive peptides that may be used to create longer acting versions of the peptides, including analogs of glucagon, glucagon-like peptide-1 (GLP-1), GLP-2, Gastric inhibitory peptide (GIP), PYY, exendin, ghrelin, gastrin, amylin, and oxyntomodulin.

This disclosure relates to methods for the prevention of the reduction of disulfide bonds in a polypeptide expressed in a recombinant host cell, comprising, following fermentation, adding selenite and/or its salts or derivatives to a harvest solution of the recombinant host cell, wherein the disulfide bond in the polypeptide remains non-reduced.

The present invention relates to the use of sulfocysteine and derivatives thereof as cell culture additives to reduce the trisulfide levels in proteins produced in cell culture.

It is described the preparation of Insulin like peptides, of chimeric Insulin like peptides and of their derivatives by the random combination of their chains A and their chains B and the pharmaceutical application of the obtained products.

Methods of refolding proteins are provided, especially cysteine-containing proteins such as insulin, proinsulin, and analogues thereof. The methods make used of tandem folding via the addition, at two different time, of two different reducing agents. A first reversible reducing agent can be added to induce folding and, at a later time, a second irreversible reducing agent can be added to prevent and/or reverse the formation of aggregates via intermolecular sulfide bonds. The methods can be used to reform a variety of cysteine-containing proteins with high yield of the native form, e.g. about 60 mol %, 70 mol %, or more.

The present invention provides a novel method for manufacturing a protein, particularly where said protein is to be coupled with another molecule. The invention further provides a method for industrial scale protein manufacturing to obtain proteins, e.g., for therapeutic purposes.

The present invention provides a method for making uncapped cysteine protein preparations, including uncapped engineered cysteine antibody preparations. The methods include, inter alia, contacting a reducing agent with engineered cysteine antibody molecules, each of the antibody molecules having at least one capped engineered cysteine residue and at least one interchain disulfide bond and reacting the reducing agent with the antibody molecules under conditions sufficient to uncap engineered cysteine residues and form cap byproducts. The method also includes removing the cap byproduct during the reduction reaction. Substantially all of the interchain disulfide bonds present in the antibody molecules prior to reduction are retained following reduction. Antibody conjugates and methods for preparing antibody conjugates using uncapped antibody preparations are also described.

The present invention pertains to methods of preventing and eliminating trisulfide bonds in proteins such as antibodies. In one embodiment, trisulfide bonds in proteins are converted to disulfide bonds as part of chromatographic purification procedures. In another embodiment, the formation of trisulfide bonds in proteins is inhibited by implementation of methods described herein during the cell culture production of such proteins. In another embodiment, monoclonal antibodies are produced by the methods described herein.

The present invention relates to an in vitro method for production of heterodimeric proteins.

Cyclized peptides having a crosslinked structure by one or more intramolecular SS bonds may be prepared by: (1-A) as to a completely protected linear peptide having two or more SH groups as functional groups on the peptide, removing protecting groups of all functional groups other than the protected SH groups in the peptide, (1-B) protecting all SH groups of the linear peptide having two or more SH groups as the functional groups on the peptide by forming a temporary SS bond, and (2) subjecting the peptide obtained by step (1-A) and step (1-B) to a folding step under oxidation and reduction conditions to obtain the cyclized peptide by re-forming an SS bond in the peptide molecule.