Naseem Farah, Khan Rizwan Hasan
Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh, 202002, India.
Protein J. 2007 Sep;26(6):415-21. doi: 10.1007/s10930-007-9081-4.
The compact intermediate of the pea lectin found to exist at pH 2.4 was treated with low (PEG-400), medium (PEG-4000) and high (PEG-20,000) molecular weight PEGs. The changes occurring in the secondary structure of the protein were monitored by CD spectropolarimetry in the far-UV range, intrinsic fluorescence was used as a probe to observe the changes in the tertiary structure which is reflected by the changes in the tryptophan environment, further ANS binding studies were made to know the extent of exposure of the hydrophobic patches which is again indicative of the overall changes occurring in the tertiary structure of the protein. It was found that the three PEGs altered the secondary as well as tertiary structure of the pH 2.4 intermediate leading to the formation of three different intermediates. The intermediates were found to have non-native secondary structure as well as non-native tertiary structure. The intermediate formed by the action of PEG-400 was due to the induction of secondary and tertiary structure while the intermediates formed under the influence of PEG-4000 and PEG-20,000 were due to loss in secondary structure and rearrangement in tertiary structure. Also the ANS binding studies showed the absence of any MG or MG-like structures formed in the folding /unfolding pathway induced by PEGs.
研究人员用低分子量（PEG - 400）、中等分子量（PEG - 4000）和高分子量（PEG - 20000）的聚乙二醇（PEG）处理了在pH 2.4条件下发现存在的豌豆凝集素紧密中间体。通过远紫外区的圆二色光谱法监测蛋白质二级结构的变化，利用内源荧光作为探针观察三级结构的变化，色氨酸环境的变化反映了三级结构的变化，还进行了8 - 苯胺基 - 1 - 萘磺酸（ANS）结合研究，以了解疏水区域的暴露程度——这同样表明了蛋白质三级结构发生的整体变化。结果发现，这三种PEG改变了pH 2.4中间体的二级和三级结构，导致形成三种不同的中间体。这些中间体具有非天然的二级结构和非天然的三级结构。由PEG - 400作用形成的中间体是二级和三级结构诱导的结果，而在PEG - 4000和PEG - 20000影响下形成的中间体是二级结构丧失和三级结构重排的结果。此外，ANS结合研究表明，在PEG诱导的折叠/去折叠途径中没有形成任何熔融球蛋白（MG）或类MG结构。
