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2 protocols using z rlrgg 7 amino 4 methyl courmarin peptide amc

1

Ubiquitin and ISG15 Substrate Assay

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Z-RLRGG-7-amino-4-methyl-courmarin (peptide-AMC) was purchased from Bachem. Ubiquitin-7amino-4-methylcourmarin (Ub-AMC) was purchased from Boston Biochem; human ISG15-7amino-4-methylcourmarin (ISG15-AMC) was purchased from Boston Biochem. Lys48 linked tetra-Ub were obtained from Boston Biochem; DLdithiothroitol (DTT) was purchased from GoldBio, and isopropyl-β-D-thiogalactopyranoside (IPTG) was purchased from GoldBio. 4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) was purchased from Fisher BioReagents. Imidazole was purchased from Acros Organics; tris(hydroxymethyl)aminomethane (Tris) was purchased from Fisher Scientific. Losartan and Sodium chloride (NaCl) were purchased from Fisher Chemical and bovine serum albumin (BSA) was purchased from Sigma Life Science.
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2

Recombinant Protein Expression and Purification

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5-Amino-2-methyl-N-[(R)-1-(1-naphthyl)ethyl]benzamide (GRL0617) was purchased from Raystar, CN; 5-(acetylamino)-2-methyl-N-[(1R)-1-(1-napthanlenyl)ethyl]-benzamide (compound 6) was purchased from MedChem Express; Z-RLRGG-7-amino-4-methyl-courmarin (peptide-AMC) was purchased from Bachem; Ubiquitin–7-amino-4-methylcourmarin (Ub-AMC) was purchased from Boston Biochem; human ISG15-7-amino-4-methylcourmarin (ISG15-AMC) was purchased from Boston Biochem; Lys6, Lys11, Lys29, Lys33, Lys48, Lys63, and linear linked di-Ub were obtained from Boston Biochem; DL-dithiothroitol (DTT) was purchased from GoldBio; Isopropyl β-D-thiogalactopyranoside (IPTG) was purchased from GoldBio; 4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) was purchased from Fisher BioReagents; Imidazole was purchased from Acros Organics; tris(hydroxymethyl)aminomethane (Tris) was purchased from Fisher Scientific; Sodium chloride (NaCl) was purchased from Fisher Chemical; Bovine serum albumin (BSA) was purchased from Sigma Life Science; Dehydrated Luria-Bertani (LB) broth was purchased from Fisher Scientific; Ampicillin was purchased from GoldBio.
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