EC Number   |
|---|
   3.4.21.75 | - |
| 3.4.21.75 | crystal structure of furin with bound decanoyl-Arg-Val-Lys-Arg-chloromethylketone inhibitor |
| 3.4.21.75 | purified enzyme furin in complexes with inhibitors N2-phenylacetyl-L-Lys-L-Lys-L-Arg-aldehyde, N2-[(3,4-dichlorophenyl)acetyl]-L-lysyl-N-[(1S)-1-(1-carbamimidoylpiperidin-4-yl)-2-oxoethyl]-L-lysinamide, N2-[(3,4-dichlorophenyl)acetyl]-L-lysyl-N-[(1S)-1-(1-carbamimidoylpiperidin-4-yl)-2-oxoethyl]-L-lysinamide, N2-(1,3-thiazol-2-yl)-L-arginyl-N-[(1S)-2-amino-2-oxo-1-(4-[[4-(trifluoromethyl)benzyl]oxy]phenyl)ethyl]-L-lysinamide, diphenyl (1-[[(benzyloxy)carbonyl]amino]-3-carbamimidamidopropyl)phosphonate, diphenyl (1-[[(benzyloxy)carbonyl]amino]-4-carbamimidamidobutyl)phosphonate, and diphenyl [2-(4-aminophenyl)-1-[[(benzyloxy)carbonyl]amino]ethyl]phosphonate, vapor diffusion method, 9.0 mg/ml glycosylated human furin in 10 mm HEPES, pH 7.5, 100 mm NaCl, and 2 mm CaCl2 is mixed with an equal volume of crystallization solution containing 100 mm MES, 200 mm K/NaH2PO4, pH 5.5-6.0, and 2m NaCl, and equilibrated against reservoir solution with 3.0-3.6m NaCl, at 20°C, crystals are soaked for 16 h in soaking solution containing 3.13 M NaCl, 100 mM Mes/NaOH, pH 5.5, 200 mM NaH2PO4, 1 mM CaCl2, and 20% DMSO supplemented with inhibitor N2-phenylacetyl-L-Lys-L-Lys-L-Arg-aldehyde (5 mM), N2-[(3,4-dichlorophenyl)acetyl]-L-lysyl-N-[(1S)-1-(1-carbamimidoylpiperidin-4-yl)-2-oxoethyl]-L-lysinamide (5 mM), N2-(4-[(Z)-[3-(cyclohexylmethyl)-2,4-dioxo-1,3-thiazolidin-5-ylidene]methyl]benzoyl)-L-lysyl-N-[(1S)-2-amino-2-oxo-1-phenylethyl]-L-lysinamide (5 mM), N2-(1,3-thiazol-2-yl)-L-arginyl-N-[(1S)-2-amino-2-oxo-1-(4-[[4-(trifluoromethyl)benzyl]oxy]phenyl)ethyl]-L-lysinamide (5 mM), diphenyl (1-[[(benzyloxy)carbonyl]amino]-3-carbamimidamidopropyl)phosphonate (5 mM), diphenyl (1-[[(benzyloxy)carbonyl]amino]-4-carbamimidamidobutyl)phosphonate (1 mM), or diphenyl [2-(4-aminophenyl)-1-[[(benzyloxy)carbonyl]amino]ethyl]phosphonate (1 mM), X-ray diffraction structure determination and analysis |
| 3.4.21.75 | purified furin in complex with a non-substrate-like small molecule inhibitor N,N'-[[(1S,3R,4R,6S)-4-carbamimidamido-6-(4-carbamimidamidoanilino)cyclohexane-1,3-diyl]bis(oxy-4,1-phenylene)]diguanidine, hexagonal crystals of unliganded furin are soaked in a 5 mM solution of inhibitor in 100 mM MES, pH 5.5, 200 mM K/NaH2PO4, 1 mM CaCl2, 10% DMSO, and 3.4 mM NaCl for 16 h, X-ray diffraction structure determination and analysis at 1.9 A resolution |
| 3.4.21.75 | purified recombinant enzyme in complex with inhibitors m-guanidinomethyl-phenylacetyl-Arg-Val-Arg-(amidomethyl)-benzamidine or phenylacetyl-Arg-Val-Arg-(amidomethyl)-benzamidine, mixing of 7.5 mg/ml enzyme with 0.290 mM m-guanidinomethyl-phenylacetyl-Arg-Val-Arg-(amidomethyl)-benzamidine and 3 mM phenylacetyl-Arg-Val-Arg-(amidomethyl)-benzamidine, respectively, and 50 mM Tris, pH 8.5, 2.8 M sodium formate and 0.015 mM Cymal-7, at 30°C, displacement of the highly potent inhibitor m-guanidinomethyl-phenylacetyl-Arg-Val-Arg-(amidomethyl)-benzamidine by competitive soaking with excessive amounts of the less potent phenylacetyl-Arg-Val-Arg-(amidomethyl)-benzamidine, X-ray diffraction structure determination and analysis at 2.7 A resolution |
| 3.4.21.75 | purified recombinant furin mutant N387D/N440D in nonglycosylated apo-form, vapor diffusion method, 400 nl of 6 mg/ml of apo-furin (aa108-574-TEV-FLAG-His6 N387D/N440D) in 10 mM HEPES, pH 7.5, 150 mM NaCl, and 5 mM CaCl2, is mixed with 400 nl of crystallization solution containing 11-13% PEG 8000, 0.11-0.16 M potassium dihydrogen phosphate, and 0.1 M HEPES, pH 7.5, at 4°C, X-ray diffraction structure determination and analysis at 1.89 A resolution. The nonglycosylated furin protein reliably forms extremely durable apo crystals that diffract to high resolution, crystals are stable at 4°C for over one year. Digestion with TEV protease for the removal of the His6 and FLAG tags is not necessary for crystallization |
| 3.4.21.75 | purified recombinant His-tagged enzyme in complexes with three peptide-derived inhibitors, vapor diffusion method, mixing of 9 mg/ml protein in 10 mM HEPES, pH 7.5, 100 mM NaCl, and 2 mM CaCl2, with an equal volume of crystallization solution containing 100 mM MES, 200 mM K/NaH2PO4, pH 5.5-6.0, 3-4 M NaCl, and 3% dimethyl sulfoxide, and equilibration against reservoir solution containing 3-4 M NaCl, 20°C, crystals are soaked for about 16 h in a soaking solution conatining 3.16 M NaCl, 100 mM MES/NaOH, pH 5.5, 200 mM Na/KH2PO4, and 1 mM CaCl2 and supplemented with 2 mM Arg-Arg-Arg-Val-Arg-4-aminomethyl-benzamidine, or 1 mM phenylacetyl-Cit-Val-Arg-4-aminomethyl-benzamidine, or 1 mM 4-aminomethyl-phenylacetyl-Arg-Tle-Arg-4-aminomethyl-benzamidine, X-ray diffraction structure determination and analysis at 1.9-2.0 A resolution |
| 3.4.21.75 | purified unliganded furin in different functional states, different Ca2+- and inhibitor (3-guanidinomethyl-phenylacetyl-Arg-Val-Arg-(4-amidomethyl)-benzamidine)-bound forms of the enzyme, crystals of human furin are grown in sitting drops mixing equal volumes of 9 mg/ml protein solution and crystallization solution containing 100 mM MES, 200 mM K/NaH2PO4, pH 5.5-6.0, 3-4 M NaCl, and 3% v/v DMSO, the reservoir contains 3-4 M NaCl, for inhibitor binding studies the crystals are soaked with inhibitor and for calcium-studies the crystals are soaked with EDTA and Ca2+, X-ray diffraction structure determination and analysis at 1.8-2.0 A resolution |
| 3.4.21.75 | triclinic crystals, space group P1, unit cell dimensions a = 93.3 A, b = 135.4 A, c = 137.8 A |
