Proteomics Approach To Trace Site-Specific Damage in Aquatic Extracellular Enzymes During Photoinactivation.
Proteomics Approach To Trace Site-Specific Damage in Aquatic Extracellular Enzymes During Photoinactivation.
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DOI：
10.1021/acs.est.7b06439
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发表时间：
2018-06
影响因子：
11.4
通讯作者：
C. Egli;E. Janssen
中科院分区：
文献类型：
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作者：
C. Egli;E. Janssen
胞外酶是地表水生态地球化学、养分和碳循环的主要驱动力。虽然光失活被认为是这些酶的主要失活过程，但基本的分子变化很少受到关注。这项研究表明，光暴露如何导致磷酸酶，氨肽酶和葡萄糖苷酶活性的生物膜样品和模型酶的快速损失。在这里，优化的蛋白质组学方法允许同时观察失活和分子变化。在三维蛋白质中产生并可视化了降解动力学的位点特异性指纹。色氨酸的氧化，发色团的目标，启动二级反应。有证据表明，酪氨酸残基作为分子内抗氧化剂，反映在减速衰减的含色氨酸的肽和增强衰减的含酪氨酸的肽。此外，随后的蛋氨酸氧化和二硫键还原有助于异质性光损伤。接近色氨酸残基解释了蛋白质结构中>95%的光损伤。溶液中氧化还原活性有机物或模型抗氧化剂的存在不仅淬灭了光失活和色氨酸氧化，而且还淬灭了所有随后的损伤。所开发的分析方法可以应用于环境科学中的其他研究问题，其中蛋白质中的位点特异性损伤是必不可少的。
Extracellular enzymes are major drivers of biogeochemical nutrient and carbon cycling in surface water. While photoinactivation is regarded as a major inactivation process of these enzymes, the underlying molecular changes have received little attention. This study demonstrates how light exposure leads to a rapid loss of phosphatase, aminopeptidase, and glucosidase activities of biofilm samples and model enzymes. Here, an optimized proteomics approach allowed simultaneous observation of inactivation and molecular changes. Site-specific fingerprints of degradation kinetics have been generated and visualized in the three-dimensional proteins. Oxidation of tryptophan, the chromophoric target, initiated secondary reactions. Evidence was obtained that tyrosine residues act as intramolecular antioxidants, reflected in decelerated decay of tryptophan-containing peptides and enhanced decay of tyrosine-containing peptides. In addition, subsequent methionine oxidation and disulfide reduction contribute to heterogeneous photodamage. The proximity to tryptophan residues explains >95% of the photodamage across the protein structures. The presence of redox active organic matter or a model antioxidant in solution quenched not only photoinactivation and tryptophan oxidation but also all subsequent damage. The developed analytical approach can be applied to other research questions in environmental sciences where site-specific damage in a protein is essential.