Neutrophil Procollagenase 蛋白

ABIN368653 产品详细信息, 供应商: Log in to see
蛋白名称
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资源
Human
蛋白类型
Native
应用范围
Screening Assay (ScA), Standard (STD)
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特异性 The full-length proenzyme consists of 447 amino acids.
交叉反应 (详细) No cross reactivity:
产品特性 Latent neutrophil procollagenase is isolated from human blood
纯度 95 %
背景 Synonyms: Neutrophil Procollagenase, Collagenase-2
Gene Name: matrix metallopeptidase 8
Gene: MMP8
分子量 75 kDa
基因ID 4317
UniProt P22894
应用备注 Degradation of extracellular Matrix, Screening and evaluation of MMP inhibitors, Antigen standard
试剂准备

Preparation and stability of solutions: APMA-solution: 40 mM p-Neutrophenyl mercuric acetate (APMA) in dimethylsulfoxide. The solution is stored at -20 °C. Peptide hydrolysis buffer: 50 mM Tris-HCl, pH 7.5, 150 mM NaCl, 5 mM CaCl 2 , 0.025 % Brij 35. The solution is stable for several weeks at 4 °C. Stock solution of peptide substrate: 100 µM solution of Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg in 20 % dimethylsulfoxide. The solution is stored at -20 °C. Stock solution of unquenched peptide: 10 µM solution of (7-methoxycoumarin-4-yl) acetyl-Pro-Leu-NH 2 (Mca-Pro-Leu) in 20 % dimethylsulfoxide. The solution is stored at -20 °C.
Activation: An aliquot of 19.5 µL Neutrophil procollagenase is mixed with 0.5 µL APMA solution and the mixture is incubated for 30 min at 37 °C.

实验流程

The activity of Neutrophil procollagenase is measured fluorimetrically with a synthetic internally quenched fluorescent substrate according to Knight et al. An excitation wavelength of 328 nm and an emission wavelength of 393 nm are set in an appropriate fluorimeter. The instrument is calibrated with the unquenched peptide Mca-Pro-Leu at a concentration corresponding to between 2 and 10 % hydrolysis of the protease substrate. Kinetic reactions are conveniently carried out in a constant volume of 2.5 mL . The substrate Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg is diluted in peptide hydrolysis buffer to a concentration of 0.8 µM and equilibrated at a temperature of 37 °C. Aliquots of 1 µL to 2 µL of the activation mixture are than added and the increase in fluorescence is recorded over a time interval between 2 and 12 min. Activity units per mL enzyme solution are calculated according to the following equation: c Mca-Pro-Leu deltaF Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg Activity U/mL = ?????.. V total F Mca-Pro-Leu v enzyme c Mca-Pro-Leu : Concentration of Mca-Pro-Leu used for calibration of the fluorimeter ( umoles/mL) F Mca-Pro-Leu : Fluorescence of Mca-Pro-Leu at the concentration c Mca-Pro-Leu used for fluorimeter calibration deltaF Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg : Change in fluorescence during peptide hydrolysis per min V : Volume of peptide hydrolysis reaction (2.5 mL ) v : Volume of added enzyme (0.001 mL to 0.002 mL ) 5. References.

限制 仅限研究用
状态 Liquid
缓冲液 50 mM Tris-HCl, pH 7.0, 200 mM NaCl, 5 mM CaCl2, 0.05 % Brij-35, 0.05 % Sodium azide.
储存液 Sodium azide
注意事项 This product contains Sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.
储存条件 -80 °C
厂商提供的图像
 image for Neutrophil Procollagenase protein (ABIN368653) Neutrophil Procollagenase protein
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