αD-芋螺毒素在东太平洋物种中的分布：以墨西哥的为例。

αD-Conotoxins in Species of the Eastern Pacific: The Case of from Mexico.

机构信息

Laboratorio de Neurofarmacología Marina, Departamento de Neurobiología Celular y Molecular, Instituto de Neurobiología, Universidad Nacional Autónoma de México, Juriquilla, Querétaro 76230, Mexico.

Departamento de Medicina Molecular y Bioprocesos, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos 62210, Mexico.

出版信息

Toxins (Basel). 2019 Jul 12;11(7):405. doi: 10.3390/toxins11070405.

DOI:10.3390/toxins11070405

PMID:31336928

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC6669827/

Abstract

snails produce venoms containing numerous peptides such as the α-conotoxins (α-CTXs), which are well-known nicotinic acetylcholine receptor (nAChR) antagonists. Thirty-eight chromatographic fractions from venom extract were isolated by RP-HPLC. The biological activities of 37 fractions (0.07 µg/µL) were assayed by two-electrode voltage clamp on human α7 nAChRs expressed in oocytes. Fractions F7 and F16 notably inhibited the response elicited by acetylcholine by 52.7 ± 15.2% and 59.6 ± 2.5%, respectively. Fraction F7 was purified, and an active peptide (F7-3) was isolated. Using a combination of Edman degradation, mass spectrometry, and RNASeq, we determined the sequence of peptide F7-3: AVKKTCIRSTOGSNWGRCCLTKMCHTLCCARSDCTCVYRSGKGHGCSCTS, with one hydroxyproline (O) and a free C-terminus. The average mass of this peptide, 10,735.54 Da, indicates that it is a homodimer of identical subunits, with 10 disulfide bonds in total. This peptide is clearly similar to αD-CTXs from species of the Indo-Pacific. Therefore, we called it αD-PiXXA. This toxin slowly and reversibly inhibited the ACh-induced response of the hα7 nAChR subtype, with an IC of 6.2 μM, and it does not affect the hα3β2 subtype at 6.5 μM.

摘要

蜗牛产生的毒液中含有许多肽，如α-CTXs（α-CTXs），它们是众所周知的烟碱型乙酰胆碱受体（nAChR）拮抗剂。通过反相高效液相色谱法（RP-HPLC）从毒液提取物中分离出 38 个色谱馏分。通过在表达于人卵母细胞中的人α7 nAChR 上进行双电极电压钳实验，测定了 37 个馏分（0.07 µg/µL）的生物活性。馏分 F7 和 F16 分别显著抑制乙酰胆碱引起的反应，抑制率分别为 52.7±15.2%和 59.6±2.5%。馏分 F7 被纯化，分离出一种活性肽（F7-3）。通过 Edman 降解、质谱和 RNAseq 的组合，我们确定了肽 F7-3 的序列：AVKKTCIRSTOGSNWGRCCLTKMCHTLCCARSDCTCVYRSGKGHGCSCTS，其中含有一个羟脯氨酸（O）和一个游离的 C 末端。该肽的平均分子量为 10735.54 Da，表明它是由相同亚基组成的同源二聚体，总共有 10 个二硫键。该肽显然与来自印度-太平洋地区物种的αD-CTXs 相似。因此，我们称之为αD-PiXXA。该毒素缓慢且可逆地抑制 ACh 诱导的 hα7 nAChR 亚型反应，IC 为 6.2 μM，在 6.5 μM 时不影响 hα3β2 亚型。