Wondrak E M, Louis J M, Mora P T, Oroszlan S
Laboratory of Molecular Virology and Carcinogenesis, NCI-Frederick Cancer Research and Development Center, MD 21702.
FEBS Lett. 1991 Mar 25;280(2):347-50. doi: 10.1016/0014-5793(91)80328-z.
Recombinant wild-type protease of human immunodeficiency virus, type 1 (HIV-1) expressed in E. coli was purified by pepstatin A affinity chromatography. An 88-fold purification was achieved giving a protease preparation with a specific enzymatic activity of approximately 3700 pmol/min/micrograms. Two proteolytically inactive HIV-1 mutant proteases (Arg-87----Lys; Asn-88----Glu) were found to bind to pepstatin A agarose, and and they were purified as the wild-type protease. A third mutant protease Arg-87----Glu) was apparently unable to bind to pepstatin A under similar conditions. Binding to pepstatin A indicates the binding ability of the substrate binding site and the ability to form dimers. These features may be used to purify and to characterize other mutated HIV-1 proteases.
