Gustafson M E, Junger K D, Foy B A, Baez J A, Bishop B F, Rangwala S H, Michener M L, Leimgruber R M, Houseman K A, Mueller R A
G.D. Searle and Co., Chesterfield, Missouri 63198, USA.
Protein Expr Purif. 1995 Aug;6(4):512-8. doi: 10.1006/prep.1995.1068.
Human immunodeficiency virus type 1 (HIV-1) protease was expressed in Escherichia coli as a fusion protein with the N-terminal sequence of IGF-2. The protein accumulated in inclusion bodies as a 40:60 mixture of unprocessed fusion protein and processed protein. A simple purification procedure was developed that yielded 30-40 mg of active protease per liter of fermentation broth with a recovery of 30-40%. The purification process involved the selective extraction of HIV-1 protease from E. coli inclusion bodies with 50% acetic acid and fractional diafiltration to remove impurities and low-molecular-weight protease-related fragments. No chromatographic steps were employed, yet the HIV-1 protease produced by this procedure was greater than 95% pure by SDS-PAGE, reverse-phase HPLC, and N-terminal sequence analysis.
