Assay methods for retroviral proteases.
A variety of assay methods for retroviral proteases have been developed in response to different experimental requirements, such as initial identification of a protease, subsequent enzymatic characterization, and high-capacity automated screening of potential inhibitors. This chapter has reviewed a number of these methods above; most have been closely tailored to match specific experimental requirements, and some of them are described in greater detail in other chapters in this volume. They include analysis of polyprotein cleavage using SDS-PAGE, analysis of the determinants of substrate cleavage using either chromogenic peptides or reversed-phase HPLC for product separation after cleavage of unmodified peptides, and the design and utilization of quenched fluoregenic peptides for use in continuous assay.
Hellen
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Department of Microbiology, State University of New York at Stonybrook 11794.
D000595:Amino Acid Sequence; D016282:Aspartic Acid Endopeptidases / Q000032:analysis* / Q000378:metabolism; D002845:Chromatography / Q000379:methods; D002863:Chromogenic Compounds; D003124:Colorimetry / Q000379:methods; D004586:Electrophoresis / Q000379:methods; D005470:Fluorometry / Q000379:methods; D006868:Hydrolysis; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000032:analysis; D010455:Peptides / Q000138:chemical synthesis; D011874:Radiometry / Q000379:methods; D012190:Retroviridae / Q000201:enzymology*; D012191:Retroviridae Proteins / Q000032:analysis* / Q000378:metabolism; D013379:Substrate Specificity
D002863:Chromogenic Compounds; D010446:Peptide Fragments; D010455:Peptides; D012191:Retroviridae Proteins; D016282:Aspartic Acid Endopeptidases