Methods for protein refolding
AssigneeBarofold, Inc.
InventorsTheodore W. RANDOLPH, John F. Carpenter, Richard J. St. John, Jonathan N. Webb
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Abstract
The present invention discloses improved methods of disaggregating protein aggregates, and refolding denatured proteins, using high pressure. In particular, the present invention provides for the use of agitation, high temperature, “stepped” depressurization, dialysis and dilution under pressure to increase the speed and extent of aggregate dissolution and protein refolding.
Claims19
A method of producing a biologically active disaggregated protein from protein aggregates comprising:
- (i) providing a protein aggregate;
- (ii) mixing said protein aggregate with a reducing agent in an amount sufficient to reduce disulfide bonds therein;
- (iii) subjecting the mixture of step (ii) to increase pressure, as compared to ambient pressure, whereby said protein aggregate dissolves;
- (iv) dialyzing the mixture under pressure, whereby said reducing agent is removed and disulfide bonds reform;
- and
- (v) removing the dissolved protein from increased pressure, whereby said protein refolds such that biological activity is retained.
12-79. (canceled)
A method of refolding denatured protein comprising:
- (i) providing a protein composition;
- (ii) subjecting the said protein composition to increased pressure, as compared to ambient pressure, and agitation, whereby said protein aggregate dissolves;
- and
- (iii) removing the protein composition from increased pressure, whereby said protein refolds such that biological activity is retained.
81-85. (canceled)
A method of inactivating a virus in a sample containing a desired protein comprising:
- (i) providing a sample containing said desired protein, said protein being in a native or non-native state;
- (ii) treating said sample to reduce or eliminate infection virus particles therein and
- (iii) subjecting said sample to a high pressure protein refolding procedure.
90-91. (canceled)
Dependencies are inferred from the claim text.