Robust substrate profiling method reveals striking differences in specificities of serum and lung fluid proteases
Robust substrate profiling method reveals striking differences in specificities of serum and lung fluid proteases
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DOI：
10.2144/000113717
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发表时间：
2011-08-01
期刊：
影响因子：
2.7
通讯作者：
Galande, Amit K.
中科院分区：
文献类型：
--
作者：
Watson, Douglas S.;Jambunathan, Kalyani;Galande, Amit K.
蛋白酶是许多疾病的候选生物标志物和治疗靶点。需要敏感和强大的技术来量化复杂生物环境中的蛋白质水解活性。我们假设组合蛋白酶底物文库可以有效地用于识别血清和支气管肺泡灌洗液（BALF）之间的异同，这两种体液对于开发靶向治疗和诊断具有重要的临床意义。我们使用一个简明的荧光探针库来绘制豚鼠血清和BALF的蛋白酶底物特异性。两种液体的蛋白水解指纹图谱的差异是惊人的：血清蛋白酶裂解含有阳离子残基和脯氨酸的底物，而BALF蛋白酶裂解含有脂肪和芳香残基的底物。值得注意的是，在人和豚鼠血清中，含有脯氨酸的底物的裂解作用主导了所有其他蛋白酶的活性。这种底物分析方法为复杂生物样品之间蛋白酶特异性的定量比较提供了基础。
Proteases are candidate biomarkers and therapeutic targets for many diseases. Sensitive and robust techniques are needed to quantify proteolytic activities within the complex biological milieu. We hypothesized that a combinatorial protease substrate library could be used effectively to identify similarities and differences between serum and bronchoalveolar lavage fluid (BALF), two body fluids that are clinically important for developing targeted therapies and diagnostics. We used a concise library of fluorogenic probes to map the protease substrate specificities of serum and BALF from guinea pigs. Differences in the proteolytic fingerprints of the two fluids were striking: serum proteases cleaved substrates containing cationic residues and proline, whereas BALF proteases cleaved substrates containing aliphatic and aromatic residues. Notably, cleavage of proline-containing substrates dominated all other protease activities in both human and guinea pig serum. This substrate profiling approach provides a foundation for quantitative comparisons of protease specificities between complex biological samples.