title: Chemical synthesis and expression of the HIV-1 protease gene in E. coli.
abstract: The 297bp HIV-1 protease gene was constructed from five discrete synthetic fragments and expressed in E. coli. A soluble protein product of 11.5 Kd was detected by immunoblotting using protease specific antisera. A quantitative assay system, utilizing a synthetic nonapeptide spanning the cleavage site between p17-p24 in the gag polyprotein, was used to measure the specific protease activity in crude extracts. The protease hydrolyzed tyrosyl-proline bonds with an approximate specific activity of 43 pmoles/min/micrograms of total protein. The chemical synthesis of the protease gene and it's expression provides a feasible method for rapid mutant analysis, important for structure-function studies and rational design of potential inhibitors.
authors.affiliation: Division of Cancer Biology and Diagnosis, National Cancer Institute, Bethesda, Maryland 20892.
authors.forename: J M
authors.initials: JM
authors.lastname: Louis
authors.forename: E M
authors.initials: EM
authors.lastname: Wondrak
authors.forename: T D
authors.initials: TD
authors.lastname: Copeland
authors.forename: C A
authors.initials: CA
authors.lastname: Smith
authors.forename: P T
authors.initials: PT
authors.lastname: Mora
authors.forename: S
authors.initials: S
authors.lastname: Oroszlan
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D004247:DNA / Q000138:chemical synthesis* / Q000235:genetics; D010450:Endopeptidases / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D005786:Gene Expression Regulation; D015683:Gene Products, gag; D016333:HIV Protease; D007700:Kinetics; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D010446:Peptide Fragments / Q000378:metabolism; D010957:Plasmids; D011498:Protein Precursors / Q000378:metabolism; D012191:Retroviridae Proteins / Q000378:metabolism; D013379:Substrate Specificity; D014169:Transformation, Bacterial
chemical_list: D015683:Gene Products, gag; D010446:Peptide Fragments; D011498:Protein Precursors; D012191:Retroviridae Proteins; D004247:DNA; D010450:Endopeptidases; D016333:HIV Protease