title: Affinity purification of the HIV-1 protease.
abstract: An inhibitor of the HIV-1 protease has been employed in the generation of a resin which allows the rapid purification of this enzyme. A peptide substrate analogue, H2N-Ser-Gln-Asn-(Phe-psi[CH2N]-Pro)-Ile-Val-Gln-OH, was coupled to agarose resin. The HIV-1 protease was expressed in E. coli and the supernatant from lysed cells was passed through the affinity resin. Active HIV-1 protease was then eluted with a buffer change to pH 10 and 2 M NaCl. Final purification to a homogeneous preparation, capable of crystallization, was achieved with hydrophobic interaction chromatography. Solutions containing HIV-1 protease bound to competitive inhibitors do not bind to the column.
authors.affiliation: Department of Molecular Biology, Merck Sharp and Dohme Research Laboratories, West Point, PA 19486.
authors.forename: J C
authors.initials: JC
authors.lastname: Heimbach
authors.forename: V M
authors.initials: VM
authors.lastname: Garsky
authors.forename: S R
authors.initials: SR
authors.lastname: Michelson
authors.forename: R A
authors.initials: RA
authors.lastname: Dixon
authors.forename: I S
authors.initials: IS
authors.lastname: Sigal
authors.forename: P L
authors.initials: PL
authors.lastname: Darke
mesh_terms: D000595:Amino Acid Sequence; D002846:Chromatography, Affinity / Q000379:methods; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D016333:HIV Protease; D015497:HIV-1 / Q000201:enzymology*; D007700:Kinetics; D008024:Ligands; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009842:Oligopeptides / Q000138:chemical synthesis / Q000494:pharmacology; D010957:Plasmids
chemical_list: D008024:Ligands; D009842:Oligopeptides; D010450:Endopeptidases; D016333:HIV Protease