脯氨酸羟基化促进芋螺毒素lt3a的氧化折叠

Effect of hydroxyproline on oxidative folding of conotoxin lt3a

毒素lt3a是从信号芋螺毒管中分离鉴定到的一种天然多肽,天然来源的lt3a的第6位脯氨酸发生了羟基化修饰。为深入研究脯氨酸羟基化的生物学意义,以lt3a中第6位脯氨酸为研究目标,采用固相化学合成的方法分别合成了不含羟基化修饰的多肽lt3a和具有羟脯氨酸的多肽lt3a[P6O],并对其氧化折叠速率、热稳定性和功能进行了研究,发现在GSSG/GSH溶液中,lt3a[P6O]的氧化折叠速率是lt3a的两倍,但两者的折叠终产率相当,进一步实验表明羟脯氨酸的存在不影响lt3a的热稳定性和对神经干复合动作电位的作用。说明羟脯氨酸促进了lt3a的氧化折叠速率,但对其生物学活性没有显著影响。该研究一方面为芋螺多肽翻译后修饰研究奠定了基础,另一方面对芋螺毒素的结构改造提供理论依据。

Conotoxin lt3a was firstly purified and identified from the crude venom of Conus litteratus . The proline in the amino acid sequence of lt3a was undergone hydroxylation modification during the process of conotoxin maturation. To investigate the effect of hydroxyproline on the folding and activity of lt3a, conotoxin lt3a [P6O] with hydroxyproline in position 6 and conotoxin lt3a without hydroxyproline were chemically synthesized and the oxidative folding rate, thermal stability and biological activity of lt3a [P6O] and lt3a were investigated. The results demonstrated that the hydroxyproline could increase the folding rate of lt3a, but had no effect on thermal stability, which was different from the situation in the collagen. It might be due to the reason that the number of Hyp in lt3a was not enough to form the helix and the stable “water bridge” structure to enhance the thermal stability. Moreover, Hyp substitution had no impact on the conotoxin activity in the case of lt3a.