Nespoulous C, Pernollet J C
Department of Plant Physiology and Biochemistry, INRA Versailles, France.
Int J Pept Protein Res. 1994 Feb;43(2):154-9. doi: 10.1111/j.1399-3011.1994.tb00516.x.
In order to investigate differences in the conformation of elicitins exhibiting different levels of activity (toxicity to tobacco plants), the environment of the tyrosyl residues in four elicitins has been compared by different spectroscopic methods (difference absorption and circular dichroism). We compared two alpha-elicitins (capsicein and parasiticein) and two beta-elicitins (beta-cryptogein and beta-cinnamomin), that are 50-100 times more toxic than the alpha-ones. Thermal difference UV spectroscopy and titration experiments clearly showed the exposure of Tyr-85 by comparison of parasiticein lacking Tyr-85 and the accessibility of its hydroxyl group to the solvent. The adjacent Tyr-87 was also suggested to be located at the surface. In beta-cryptogein, beta-cinnamomin and capsicein the pK was measured at between 10.5 and 10.8, while in parasiticein it is higher (11.5) owing to a difference in the local environment. Thermal difference UV spectroscopy showed one more exposed tyrosine in beta-elicitins than in alpha-ones. This difference was attributed to Tyr-12, considering the more hydrophilic characteristic of the sequence around residue 13 in beta-elicitins and the role of this region in the toxicity. However, no difference in titration behaviour was noted among elicitins concerning Tyr-12. The other two tyrosines also presented an abnormal pK of titration (> 12). In all elicitins Tyr-47 was probably exposed, while Tyr-33 was probably buried and not titrated, except in beta-cinnamomin at very alkaline pH.
为了研究具有不同活性水平（对烟草植物的毒性）的激发素在构象上的差异，通过不同的光谱方法（差分吸收和圆二色性）比较了四种激发素中酪氨酸残基的环境。我们比较了两种α-激发素（辣椒激发素和寄生疫霉激发素）和两种β-激发素（β-隐地蛋白和β-肉桂霉素），它们的毒性比α-激发素高50-100倍。通过比较缺乏酪氨酸-85的寄生疫霉及其羟基对溶剂的可及性，热差紫外光谱和滴定实验清楚地表明了酪氨酸-85的暴露。相邻的酪氨酸-87也被认为位于表面。在β-隐地蛋白、β-肉桂霉素和辣椒激发素中，测得的pK在10.5至10.8之间，而在寄生疫霉中由于局部环境的差异其pK较高（11.5）。热差紫外光谱显示，β-激发素中比α-激发素中多一个暴露的酪氨酸。考虑到β-激发素中残基13周围序列的亲水性更强以及该区域在毒性中的作用，这种差异归因于酪氨酸-12。然而，在激发素之间关于酪氨酸-12的滴定行为没有差异。其他两个酪氨酸的滴定pK也异常（>12）。在所有激发素中，酪氨酸-47可能是暴露的，而酪氨酸-33可能被掩埋且未被滴定，除了在非常碱性的pH条件下的β-肉桂霉素。
