Department of Chemistry and Biotechnology, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan.
J Biosci Bioeng. 2011 Jan;111(1):10-5. doi: 10.1016/j.jbiosc.2010.08.016. Epub 2010 Sep 19.
This paper presents the synergistic enhancement of the refolding yield of denatured and reduced lysozyme by using detergents as aggregation inhibitors and water-miscible organic cosolvents as modulators for the detergents. Adding only cetyltrimethylammonium bromide (CTAB) led to a slight increase in the refolding yield (up to 13%). Further addition of dimethylsulfoxide (DMSO) with CTAB drastically increased the refolding yield up to 35%, a value which was higher than the simple sum of the refolding yields in the presence of only CTAB or DMSO. The synergistic enhancement was also observed in the coexistence of other detergents, such as polyethylene glycol monooleyl ether (n = 50) and N-tetradecyl-N,N-dimethyl-3-ammonio-1-propanesulfonate, and cosolvents, such as N,N-dimethylformamide and N,N-dimethylacetamide. Experimental data and a kinetic analysis revealed the guideline for selecting a couple of additives; detergents which can adequately inhibit the aggregation of proteins by binding to hydrophobic surfaces of refolding intermediates should be employed as an aggregation inhibitor, and cosolvents which can properly prevent both protein-protein and protein-detergent interactions act as effective modulators for the aggregation inhibitor, resulting in a desirable balance between folding and aggregation rates.
本文提出了一种协同增强方法，即在作为聚集抑制剂的去污剂和作为去污剂调节剂的水溶性有机溶剂中，可协同增强变性和还原溶菌酶的复性产率。仅添加十六烷基三甲基溴化铵（CTAB）可使复性产率略有增加（最高可达 13%）。进一步添加 CTAB 的二甲亚砜（DMSO）可使复性产率急剧增加至 35%，这一数值高于仅存在 CTAB 或 DMSO 时的复性产率之和。在共存其他去污剂（如聚乙二醇单油醚（n = 50）和 N-十四烷基-N，N-二甲基-3-氨丙基磺酸钠）和共溶剂（如 N，N-二甲基甲酰胺和 N，N-二甲基乙酰胺）时，也观察到协同增强作用。实验数据和动力学分析揭示了选择一对添加剂的指导原则；应选择能够通过与复性中间体的疏水表面结合来充分抑制蛋白质聚集的去污剂作为聚集抑制剂，并且能够适当防止蛋白质-蛋白质和蛋白质-去污剂相互作用的共溶剂作为聚集抑制剂的有效调节剂，从而在折叠和聚集速率之间达到理想的平衡。
