Guatimosim S C, Kalapothakis E, Diniz C R, Chávez-Olórtegui C
Departamento de Bioquímica e Imunologia, ICB, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil.
Toxicon. 2000 Jan;38(1):113-21. doi: 10.1016/s0041-0101(99)00138-5.
An immunogenic nontoxic protein (TsNTxP) was purified from the venom of the scorpion Tityus serrulatus (Ts). This peptide is composed of 63 amino acid residues with a high degree of structural homology with the toxins isolated from Ts. The nucleotide sequence of the gene that encodes TsNTxP was obtained and also showed a high degree of similarity with genes encoding Tityus toxins [Guatimosim, S.C.F., Prado, V.F., Diniz, C.R., Chávez-Olórtegui, C.. Kalapothakis, E., 1999. Molecular cloning and genomic analysis of TsNTxP; an immunogenic protein from Tityus serrulatus scorpion venom. Toxicon 37, 507-517]. In the present study the TsNTxP gene was expressed in E. coli BL21DE3 cells as a fusion protein with maltose-binding protein. The recombinant protein (TsNTxPrec) was purified by affinity chromatography and used as an immunogen in rabbits. The antigenic specificity of anti-TsNTxPrec antibodies was compared by an enzyme-linked immunosorbent assay using TsNTxP, TstFG50 (the fraction of Ts venom that represents most of the toxicity of the crude venom) and the crude venom, to coat microtitration plates. Anti-TsNTxPrec antibodies had a comparable high cross-reactivity for all antigens tested. Concentrations of Ts venom equivalent to 20 LD50 were effectively neutralized by 1 ml of the anti-TsNTxPrec serum. This result provides basic data for the use of such recombinant scorpion protein as an immunogen in the development of antivenoms for clinical use.
从锯脂鲤蝎（Tityus serrulatus，Ts）毒液中纯化出一种具有免疫原性的无毒蛋白（TsNTxP）。该肽由63个氨基酸残基组成，与从Ts分离出的毒素具有高度的结构同源性。获得了编码TsNTxP的基因的核苷酸序列，其与编码锯脂鲤毒素的基因也显示出高度相似性[瓜蒂莫西姆，S.C.F.，普拉多，V.F.，迪尼兹，C.R.，查韦斯 - 奥洛尔特吉，C.，卡拉波塔基斯，E.，1999年。TsNTxP的分子克隆和基因组分析；锯脂鲤蝎毒液中的一种免疫原性蛋白。《毒素学》37卷，507 - 517页]。在本研究中，TsNTxP基因在大肠杆菌BL21DE3细胞中作为与麦芽糖结合蛋白的融合蛋白表达。重组蛋白（TsNTxPrec）通过亲和层析纯化，并用作兔的免疫原。使用TsNTxP、TstFG50（代表粗毒液大部分毒性的Ts毒液组分）和粗毒液包被微量滴定板，通过酶联免疫吸附测定法比较抗TsNTxPrec抗体的抗原特异性。抗TsNTxPrec抗体对所有测试抗原具有相当高的交叉反应性。1毫升抗TsNTxPrec血清可有效中和相当于20 LD50的Ts毒液浓度。该结果为使用这种重组蝎蛋白作为免疫原开发临床用抗蛇毒血清提供了基础数据。
