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J Protein Chem. 1993 Jun;12(3):323-7.

Large scale purification and refolding of HIV-1 protease from Escherichia coli inclusion bodies.

Author information

1
Upjohn Company, Kalamazoo, Michigan 49001.

Abstract

The protease encoded by the human immunodeficiency virus type 1 (HIV-1) was engineered in Escherichia coli as a construct in which the natural 99-residue polypeptide was preceded by an NH2-terminal methionine initiator. Inclusion bodies harboring the recombinant HIV-1 protease were dissolved in 50% acetic acid and the solution was subjected to gel filtration on a column of Sephadex G-75. The protein, eluted in the second of two peaks, migrated in SDS-PAGE as a single sharp band of M(r) approximately 10,000. The purified HIV-1 protease was refolded into an active enzyme by diluting a solution of the protein in 50% acetic acid with 25 volumes of buffer at pH 5.5. This method of purification, which has also been applied to the purification of HIV-2 protease, provides a single-step procedure to produce 100 mg quantities of fully active enzyme.

PMID:
8397790
DOI:
10.1007/bf01028194
[Indexed for MEDLINE]

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