Extracellular proteins are crucial for the formation and structural maintenance of anammox granules. However, the influence of extracellular polymeric substances (EPS) extraction methods on the secondary structure of extracellular proteins has not been studied fully. In this work, ten EPS extraction methods were applied to anammox granules and compared systematically. The EPS extraction methods were then applied to bovine serum albumin (BSA), a representative of extracellular proteins, to investigate their influence on protein secondary structure by circular dichroism (CD) spectroscopy. The ten EPS extraction methods yielded EPS concentration varying from 5.2 ± 0.1 to 84.8 ± 2.0 mg/g volatile suspended solid (VSS). Tween-20 treatment resulted in the most similar CD spectrum and protein secondary structure to the control. Treatment 0.5% Tween-20 for 4 h was found to be the best condition to preserve protein secondary structure while achieving a high EPS yield (44.4 ± 1.4 mg/g VSS) and limited cell lysis for anammox granules. This study provides an optimized protocol to extract granular EPS when the role of the secondary structure of extracellular proteins in regulating aggregation, adsorption and biofilm formation of microbes is to be elucidated.

细胞外蛋白对于厌氧氨氧化颗粒的形成和结构维持至关重要。但是，尚未充分研究细胞外聚合物（EPS）提取方法对细胞外蛋白二级结构的影响。在这项工作中，将十种EPS提取方法应用于厌氧氨氧化颗粒并进行系统比较。然后将EPS提取方法应用于牛血清白蛋白（BSA）（一种细胞外蛋白质），以通过圆二色性（CD）光谱研究其对蛋白质二级结构的影响。十种EPS提取方法产生的EPS浓度在5.2±0.1至84.8±2.0 mg / g挥发性悬浮固体（VSS）之间。Tween-20处理产生与对照最相似的CD光谱和蛋白质二级结构。治疗0。发现5％Tween-20持续4 h是保持蛋白质二级结构，同时实现高EPS产量（44.4±1.4 mg / g VSS）和厌氧菌颗粒有限的细胞裂解的最佳条件。当要阐明细胞外蛋白的二级结构在调节微生物的聚集，吸附和生物膜形成中的作用时，本研究提供了提取粒状EPS的优化方案。