Malcolm Andrew S, Dexter Annette F, Middelberg Anton P J
Centre for Biomolecular Engineering, University of Queensland, St Lucia, QLD 4072, Australia.
Langmuir. 2006 Oct 10;22(21):8897-905. doi: 10.1021/la060565u.
We present the first characterization of the mechanical properties of lysozyme films formed by self-assembly at the air-water interface using the Cambridge interfacial tensiometer (CIT), an apparatus capable of subjecting protein films to a much higher level of extensional strain than traditional dilatational techniques. CIT analysis, which is insensitive to surface pressure, provides a direct measure of the extensional stress-strain behavior of an interfacial film without the need to assume a mechanical model (e.g., viscoelastic), and without requiring difficult-to-test assumptions regarding low-strain material linearity. This testing method has revealed that the bulk solution pH from which assembly of an interfacial lysozyme film occurs influences the mechanical properties of the film more significantly than is suggested by the observed differences in elastic moduli or surface pressure. We have also identified a previously undescribed pH dependency in the effect of solution ionic strength on the mechanical strength of the lysozyme films formed at the air-water interface. Increasing solution ionic strength was found to increase lysozyme film strength when assembly occurred at pH 7, but it caused a decrease in film strength at pH 11, close to the pI of lysozyme. This result is discussed in terms of the significant contribution made to protein film strength by both electrostatic interactions and the hydrophobic effect. Washout experiments to remove protein from the bulk phase have shown that a small percentage of the interfacially adsorbed lysozyme molecules are reversibly adsorbed. Finally, the washout tests have probed the role played by additional adsorption to the fresh interface formed by the application of a large strain to the lysozyme film and have suggested the movement of reversibly bound lysozyme molecules from a subinterfacial layer to the interface.
我们首次使用剑桥界面张力仪（CIT）对在空气-水界面通过自组装形成的溶菌酶薄膜的力学性能进行了表征。该仪器能够使蛋白质薄膜承受比传统膨胀技术更高水平的拉伸应变。CIT分析对表面压力不敏感，无需假设机械模型（如粘弹性），也无需对低应变材料线性做出难以验证的假设，就能直接测量界面薄膜的拉伸应力-应变行为。这种测试方法表明，形成界面溶菌酶薄膜的本体溶液pH值对薄膜力学性能的影响比弹性模量或表面压力的观测差异所表明的更为显著。我们还发现了溶液离子强度对在空气-水界面形成的溶菌酶薄膜机械强度的影响中，存在一种此前未描述的pH依赖性。当在pH 7时进行组装，发现增加溶液离子强度会提高溶菌酶薄膜强度，但在接近溶菌酶pI的pH 11时，却会导致薄膜强度降低。从静电相互作用和疏水效应对蛋白质薄膜强度的重大贡献方面对该结果进行了讨论。从本体相中去除蛋白质的洗脱实验表明，一小部分界面吸附的溶菌酶分子是可逆吸附的。最后，洗脱试验探究了对溶菌酶薄膜施加较大应变形成的新界面上额外吸附所起的作用，并表明可逆结合的溶菌酶分子从亚界面层向界面移动。
