Fields Gregg B
Department of Chemistry and Biochemistry, Florida Atlantic University, Boca Raton, FL, USA.
Methods Mol Biol. 2010;622:393-433. doi: 10.1007/978-1-60327-299-5_24.
A continuous assay method, such as the one that utilizes an increase in fluorescence upon hydrolysis, allows for rapid and convenient kinetic evaluation of proteases. To better understand MMP behaviors and to aid in the design of MMP inhibitors, a variety of sequence specificity, phage display, and combinatorial chemistry studies have been performed. Results of these studies have been valuable for defining the differences in MMPs and for creating quenched fluorescent substrates that utilize fluorescence resonance energy transfer (FRET)/intramolecular fluorescence energy transfer (IFET). FRET triple-helical substrates have been constructed to examine the collagenolytic activity of MMP family members. The present chapter provides an overview of MMP and related FRET substrates and describes how to construct and utilize these substrates.
一种连续检测方法，例如利用水解时荧光增强的方法，可实现对蛋白酶的快速便捷的动力学评估。为了更好地理解基质金属蛋白酶（MMP）的行为并辅助MMP抑制剂的设计，已经开展了各种序列特异性、噬菌体展示和组合化学研究。这些研究结果对于界定MMP之间的差异以及创建利用荧光共振能量转移（FRET）/分子内荧光能量转移（IFET）的淬灭荧光底物很有价值。已经构建了FRET三螺旋底物来检测MMP家族成员的胶原olytic活性。本章概述了MMP及相关的FRET底物，并描述了如何构建和利用这些底物。
