Macromolecular Substrates For Enzymes

酶活性的测量是疾病过程诊断和基础研究的重要工具。我们正在寻求开发新的酶底物，以提高测量酶活性的能力或特异性。我们发现，将小的显色和荧光底物与聚合物分子连接可以作为制备大分子大小底物的方法。临床化学2001；47:215-222描述了初步结果。底物的分子大小主要由所附聚合物的大小决定，因此它是可调节的。这使得底物大小可以作为一个自变量来分析，并且它可以作为评估酶活性位点的空间位阻的一种手段。大型合成底物可作为天然底物的较好模型，如生理蛋白酶的蛋白质底物和淀粉酶的大型淀粉底物。使用大底物似乎可以区分游离蛋白酶分子和那些与抑制剂-2-巨球蛋白络合的蛋白酶分子。因此，这些底物似乎对更准确地测量血浆蛋白酶（如凝血酶）的功能活性具有实用价值。

Measurements of enzyme activity serve as important tools for the diagnosis of disease processes and for basic research. We are seeking to develop new substrates for enzymes that improve the ability to measure or the specificity of measuring enzyme activity. We have found that linking small chromogenic and fluorogenic substrates to polymer molecules serves as an approach to prepare substrates with large molecular size. Initial results are described in Clinical Chemistry 2001;47:215-222. The molecular size of the substrates is determined primarily by the size of the attached polymer so that it is adjustable. This permits substrate size to be analyzed as an independent variable, and it serves as a means of assessing the steric hindrance of the active sites of enzymes. The large synthetic substrates serve as better models of natural substrates that are of large molecular size such as the protein substrates of physiological proteinases and the large starch substrates of amylase. Use of large substrates appears to distinguish free protease molecules from those complexed with the inhibitor alpha-2-macroglobulin. Thus, these substrates appear to have utility for the more accurate measurement of the functional activity of plasma proteinases such as thrombin.