A substrate-optimized electrophoretic mobility shift assay for ADAM12

作者：Kotzsch Alexander^*; Skovgaard Tine; Buus Uwe; Andersen Simon; Devkota Kanchan; Berthelsen Jens

来源：Analytical Biochemistry, 2014, 452: 34-42.

DOI：10.1016/j.ab.2014.02.008

• Metalloprotease
• ADAM12
• Fluorimetric assay
• Mobility shift assay
• Small-molecule inhibitor
• Peptide array

ADAM12 belongs to the A disintegrin and metalloprotease (ADAM) family of secreted sheddases activating extracellular growth factors such as epidermal growth factor receptor (EGFR) ligands and tumor necrosis factor-alpha (TNF-alpha). ADAM proteases, most notably ADAM17 (TNF-alpha-converting enzyme), have long been investigated as pharmaceutical drug targets; however, due to lack of potency and in vivo side effects, none of the small-molecule inhibitors discovered so far has made it beyond clinical testing. Ongoing research on novel selective inhibitors of ADAMs requires reliable biochemical assays to validate molecular probes from large-scale screening efforts. Here we describe an electrophoretic mobility shift assay for ADAM12 based on the identification of an optimized peptide substrate that is characterized by excellent performance and reproducibility.

• 出版日期2014-5-1