Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Lab for Marine Drug of Haikou, Hainan University, Haikou, Hainan 570228, China.
Mar Drugs. 2013 Sep 17;11(9):3537-53. doi: 10.3390/md11093537.
Alpha6beta2 nicotinic acetylcholine receptors (nAChRs) are potential therapeutic targets for the treatment of several neuropsychiatric diseases, including addiction and Parkinson's disease. Alpha-conotoxin (α-CTx) TxIB is a uniquely selective ligand, which blocks α6/α3β2β3 nAChRs only, but does not block the other subtypes. Therefore, α-CTx TxIB is a valuable therapeutic candidate peptide. Synthesizing enough α-CTx TxIB with high yield production is required for conducting wide-range testing of its potential medicinal applications. The current study optimized the cleavage of synthesized α-CTx TxIB resin-bounded peptide and folding of the cleaved linear peptide. Key parameters influencing cleavage and oxidative folding of α-CTx TxIB were examined, such as buffer, redox agents, pH, salt, co-solvent and temperature. Twelve conditions were used for cleavage optimization. Fifty-four kinds of one-step oxidative solution were used to assess their effects on each α-CTx TxIB isomers' yield. The result indicated that co-solvent choices were particularly important. Completely oxidative folding of globular isomer was achieved when the NH₄HCO₃ or Tris-HCl folding buffer at 4 °C contained 40% of co-solvent DMSO, and GSH:GSSG (2:1) or GSH only with pH 8~8.7.
α6β2 型烟碱型乙酰胆碱受体 (nAChRs) 是治疗多种神经精神疾病的潜在治疗靶点，包括成瘾和帕金森病。α-芋螺毒素 (α-CTx) TxIB 是一种独特选择性配体，仅阻断 α6/α3β2β3 nAChRs，而不阻断其他亚型。因此，α-CTx TxIB 是一种有价值的治疗候选肽。为了广泛测试其潜在的药用应用，需要用高产率生产足够量的 α-CTx TxIB。本研究优化了合成的 α-CTx TxIB 树脂结合肽的切割和切割线性肽的折叠。考察了影响 α-CTx TxIB 切割和氧化折叠的关键参数，如缓冲液、氧化还原试剂、pH 值、盐、共溶剂和温度。对 12 种条件进行了切割优化。评估了 54 种一步氧化溶液对每种 α-CTx TxIB 异构体产率的影响。结果表明，共溶剂的选择尤为重要。当 4°C 时 NH₄HCO₃ 或 Tris-HCl 折叠缓冲液中含有 40%的共溶剂 DMSO 时，球状异构体可完全进行氧化折叠，并且 GSH:GSSG（2:1）或仅 GSH 在 pH8~8.7 时也可进行折叠。
