Department of Crop and Soil Science , Oregon State University , Corvallis , Oregon 97331 , United States.
Environmental Molecular Science Laboratory , Pacific Northwest National Laboratory , Richland , Washington 99352 , United States.
Environ Sci Technol. 2019 Mar 19;53(6):3018-3026. doi: 10.1021/acs.est.8b05583. Epub 2019 Feb 28.
We investigated the extent to which contact with mineral surfaces affected the molecular integrity of a model protein, with an emphasis on identifying the mechanisms (hydrolysis, oxidation) and conditions leading to protein alteration. To this end, we studied the ability of four mineral surface archetypes (negatively charged, positively charged, neutral, redox-active) to abiotically fragment a well-characterized protein (GB1) as a function of pH and contact time. GB1 was exposed to the soil minerals montmorillonite, goethite, kaolinite, and birnessite at pH 5 and pH 7 for 1, 8, 24, and 168 h and the supernatant was screened for peptide fragments using Tandem Mass Spectrometry. To distinguish between products of oxidative and hydrolytic cleavage, we combined results from the SEQUEST algorithm, which identifies protein fragments that were cleaved hydrolytically, with the output of a deconvolution algorithm (DECON-Routine) designed to identify oxidation fragments. All four minerals were able to induce protein cleavage. Manganese oxide was effective at both hydrolytic and oxidative cleavage. The fact that phyllosilicates-which are not redox active-induced oxidative cleavage indicates that surfaces acted as catalysts and not as reactants. Our results extend previous observations of proteolytic capabilities in soil minerals to the groups of phyllosilicates and Fe-oxides. We identified structural regions of the protein with particularly high susceptibility to cleavage (loops and β strands) as well as regions that were entirely unaffected (α helix).
我们研究了与矿物表面接触对模型蛋白质分子完整性的影响程度，重点是确定导致蛋白质改变的机制（水解、氧化）和条件。为此，我们研究了四种矿物表面原型（带负电荷、带正电荷、中性、氧化还原活性）在 pH 值和接触时间的作用下，使一种经过充分表征的蛋白质（GB1）非生物断裂的能力。GB1 在 pH 5 和 pH 7 下与蒙脱石、针铁矿、高岭石和水钠锰矿接触 1、8、24 和 168 小时，并用串联质谱法对上清液中的肽片段进行筛选。为了区分氧化和水解裂解的产物，我们将 SEQUEST 算法的结果（该算法识别出水解裂解的蛋白质片段）与专门用于识别氧化片段的解卷积算法（DECON-Routine）的输出结果结合起来。四种矿物都能诱导蛋白质裂解。氧化锰既能进行水解又能进行氧化裂解。层状硅酸盐——不是氧化还原活性物质——能诱导氧化裂解，这表明表面起到了催化剂的作用，而不是反应物。我们的研究结果将土壤矿物中的蛋白水解能力扩展到了层状硅酸盐和铁氧化物这两组矿物。我们确定了对切割特别敏感的蛋白质结构区域（环和β链）以及完全不受影响的区域（α螺旋）。
