Collet Beatrice Y M, Nagashima Tadamichi, Yu Marvin S, Pohl Nicola L B
Department of Chemistry and the Plant Sciences Institute, Gilman Hall, Iowa State University, Ames, IA, 50011-3111.
J Fluor Chem. 2009 Nov 1;130(11):1042-1048. doi: 10.1016/j.jfluchem.2009.09.005.
As ever more protease sequences are uncovered through genome sequencing projects, efficient parallel methods to discover the potential substrates of these proteases becomes crucial. Herein we describe the first use of fluorous-based microarrays to probe peptide sequences and begin to define the scope and limitations of fluorous microarray technologies for the screening of proteases. Comparison of a series of serine proteases showed that their ability to cleave peptide substrates in solution was maintained upon immobilization of these substrates onto fluorous-coated glass slides. The fluorous surface did not serve to significantly inactivate the enzymes. However, addition of hydrophilic components to the peptide sequences could induce lower rates of substrate cleavage with enzymes such as chymotrypsin with affinities to hydrophobic moieties. This work represents the first step to creating robust protease screening platforms using noncovalent microarray interface that can easily incorporate a range of compounds on the same slide.
随着通过基因组测序项目发现越来越多的蛋白酶序列，开发高效的并行方法来发现这些蛋白酶的潜在底物变得至关重要。在此，我们描述了首次使用基于氟的微阵列来探测肽序列，并开始确定用于蛋白酶筛选的氟微阵列技术的范围和局限性。对一系列丝氨酸蛋白酶的比较表明，当将这些底物固定在氟涂层载玻片上时，它们在溶液中切割肽底物的能力得以保持。氟表面并没有显著使酶失活。然而，向肽序列中添加亲水性成分可能会导致诸如对疏水部分具有亲和力的胰凝乳蛋白酶等酶的底物切割速率降低。这项工作代表了使用非共价微阵列界面创建强大的蛋白酶筛选平台的第一步，该界面可以轻松地在同一张载玻片上整合一系列化合物。
