Del Vecchio Pompea, Graziano Giuseppe, Granata Vincenzo, Farias Tiziana, Barone Guido, Mandrich Luigi, Rossi Mosè, Manco Giuseppe
Department of Chemistry, University of Naples Federico II, Italy.
Biochemistry. 2004 Nov 23;43(46):14637-43. doi: 10.1021/bi048344f.
The stability of acetyl-esterase, Aes, from Escherichia coli against the denaturing action of urea and guanidine hydrochloride, GuHCl, has been investigated by means of circular dichroism and fluorescence measurements. The urea-induced unfolding curves show a single inflection point at 6.2 M urea, whereas the GuHCl-induced curves show two inflection points at 1.4 and 3.1 M GuHCl. The unfolding process is reversible with both urea and GuHCl. These results, together with similar experimental data on the mutant form V20D-Aes, suggest the presence of two domains in the Aes structure, which unfold more or less independently depending on the denaturant used. This is also supported by a 3D model obtained by homology modeling using the structure of brefeldine as a template. The effect of NaCl on the urea-induced unfolding curves of the enzyme has also been investigated.
通过圆二色性和荧光测量手段，研究了来自大肠杆菌的乙酰酯酶（Aes）对尿素和盐酸胍（GuHCl）变性作用的稳定性。尿素诱导的去折叠曲线在6.2 M尿素处显示一个单一拐点，而GuHCl诱导的曲线在1.4 M和3.1 M GuHCl处显示两个拐点。去折叠过程对尿素和GuHCl都是可逆的。这些结果，连同关于突变体形式V20D - Aes的类似实验数据，表明Aes结构中存在两个结构域，它们根据所用变性剂的不同或多或少独立地去折叠。这也得到了以布雷菲德菌素结构为模板通过同源建模获得的三维模型的支持。还研究了NaCl对该酶尿素诱导的去折叠曲线的影响。
