The KEX2-encoded endoprotease was overproduced in yeast several hundred-fold and further purified to achieve a 10,000-fold enrichment in specific activity. The enzyme was (i) membrane-bound, but solubilized by detergents; (ii) able to cleave peptide substrates at both Lys-Arg and Arg-Arg sites; (iii) inhibited by EDTA and EGTA (but not o-phenanthroline), but fully reactivated by Ca2+; (iv) unaffected by 5-10 mM phenylmethylsulfonyl fluoride, N alpha-(ptosyl)lysine chloromethyl ketone, or L-1-tosylamido-2-phenylethyl chloromethyl ketone, but inactivated by 1-2 microM Ala-Lys-Arg-chloromethyl ketone; (v) labeled specifically by 125I-labeled Tyr-Ala-Lys-Arg-chloromethyl ketone; and (vi) resistant to trans-epoxysuccinate compounds (which inactivate thiol proteases), but inactivated by diisopropyl fluorophosphate (a diagnostic serine protease inhibitor). Mutant enzyme molecules lacking as many as 200 C-terminal residues still retained Ca2+-dependent protease activity and were labeled by 125I-labeled Tyr-Ala-Lys-Arg-chloromethyl ketone.
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Evidence ID | Analyze ID | Gene/Complex | Systematic Name/Complex Accession | Qualifier | Gene Ontology Term ID | Gene Ontology Term | Aspect | Annotation Extension | Evidence | Method | Source | Assigned On | Reference |
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Evidence ID | Analyze ID | Gene | Gene Systematic Name | Phenotype | Experiment Type | Experiment Type Category | Mutant Information | Strain Background | Chemical | Details | Reference |
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Evidence ID | Analyze ID | Gene | Gene Systematic Name | Disease Ontology Term | Disease Ontology Term ID | Qualifier | Evidence | Method | Source | Assigned On | Reference |
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Evidence ID | Analyze ID | Regulator | Regulator Systematic Name | Target | Target Systematic Name | Direction | Regulation of | Happens During | Regulator Type | Direction | Regulation Of | Happens During | Method | Evidence | Strain Background | Reference |
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Site | Modification | Modifier | Source | Reference |
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Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Allele | Assay | Annotation | Action | Phenotype | SGA score | P-value | Source | Reference | Note |
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Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Assay | Annotation | Action | Modification | Source | Reference | Note |
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Complement ID | Locus ID | Gene | Species | Gene ID | Strain background | Direction | Details | Source | Reference |
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Evidence ID | Analyze ID | Dataset | Description | Keywords | Number of Conditions | Reference |
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