Legros C, Feyfant E, Sampieri F, Rochat H, Bougis P E, Martin-Eauclaire M F
Laboratoire de Biochimie, Ingénierie des Protéines, UMR 6560 du Centre National de la Recherche Scientifique, Institut Fédératif Jean Roche, Faculté de Médecine Nord, Marseille, France.
FEBS Lett. 1997 Nov 3;417(1):123-9. doi: 10.1016/s0014-5793(97)01177-0.
A cDNA encoding a short polypeptide blocker of K+ channels, kaliotoxin 2 (KTX2), from the venom of the North African scorpion Androctonus australis was expressed in the periplasmic space of Escherichia coli. KTX2 was produced as a fusion protein with the maltose binding protein followed by the recognition site for factor Xa or enterokinase preceding the first amino acid residue of the toxin. The fully refolded recombinant KTX2 (rKTX2) was obtained (0.15-0.30 mg/l of culture) and was indistinguishable from the native toxin according to chemical and biological criteria. An N-extended analogue of KTX2 exhibiting three additional residues was also expressed. This analogue had 1000-fold less affinity for the 125I-kaliotoxin binding site on rat brain synaptosomes than KTX2. Conformational models of KTX2 and its mutant were designed by amino acid replacement using the structure of agitoxin 2 from Leiurus quinquestriatus as template, to try to understand the decrease in affinity for the receptor.
从北非蝎子澳毒蝎（Androctonus australis）毒液中编码钾离子通道短多肽阻断剂卡利毒素2（KTX2）的互补DNA（cDNA）在大肠杆菌的周质空间中表达。KTX2作为与麦芽糖结合蛋白的融合蛋白产生，在毒素的第一个氨基酸残基之前有凝血因子Xa或肠激酶的识别位点。获得了完全重折叠的重组KTX2（rKTX2）（0.15 - 0.30毫克/升培养物），根据化学和生物学标准，它与天然毒素没有区别。还表达了一种具有三个额外残基的KTX2的N端延伸类似物。该类似物对大鼠脑突触体上125I - 卡利毒素结合位点的亲和力比KTX2低1000倍。以五条纹肥尾蝎（Leiurus quinquestriatus）的阿吉毒素2的结构为模板，通过氨基酸置换设计了KTX2及其突变体的构象模型，以试图理解对受体亲和力降低的原因。
