Department of Biophysics, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran.
Int J Biol Macromol. 2011 Nov 1;49(4):652-6. doi: 10.1016/j.ijbiomac.2011.06.025. Epub 2011 Jul 1.
The stability of enzymes with no reduction in their catalytic activity still remains a critical issue in industrial applications. Naturally occurring osmolytes are commonly used as protein stabilizers. In this study we have investigated the effects of sorbitol and trehalose on the structural stability and activity of Pseudomonas cepacia lipase (PCL), using UV-visible, circular dichroism (CD) and fluorescence spectroscopy. Surface plasmon resonance (SPR) technique was used to trace changes in the refractive index and dielectric constant of the environment. The results revealed that catalytic activity and intrinsic fluorescence intensity of PCL increased in the presence of both osmolytes. Far-UV CD spectra indicated that the protein has undergone some conformational changes upon interacting with these osmolytes. Increasing the concentration of sorbitol led to changes in the refractive index and consequently the dielectric constant of environment; whereas in the case of trehalose, such changes were not significant. Unfavorable interactions of trehalose with protein surface induced higher preferential exclusion from the enzyme-water interface than that of sorbitol. Results of this report could give further insights about the stabilization mechanism of osmolytes.
在工业应用中，保持酶的稳定性而不降低其催化活性仍然是一个关键问题。天然存在的渗透剂通常被用作蛋白质稳定剂。在这项研究中，我们使用紫外-可见分光光度法、圆二色性（CD）和荧光光谱法研究了山梨醇和海藻糖对假单胞菌脂肪酶（PCL）结构稳定性和活性的影响。表面等离子体共振（SPR）技术用于跟踪环境折射率和介电常数的变化。结果表明，在存在两种渗透剂的情况下，PCL 的催化活性和本征荧光强度均增加。远紫外 CD 光谱表明，蛋白质在与这些渗透剂相互作用时发生了一些构象变化。山梨醇浓度的增加导致环境折射率和介电常数发生变化；而在海藻糖的情况下，这种变化并不显著。海藻糖与蛋白质表面的不利相互作用导致其从酶-水界面的优先排斥程度高于山梨醇。本报告的结果可以进一步了解渗透剂的稳定机制。
