A kind of toxin protein and its purification method

Abstract

The invention provides a toxin protein which is separated and purified from latrodectus tredecimguttatus eggs, wherein the N-terminal sequence is Glu-Ser-Ile-Gln-Thr-Ser-Thr-Tyr-Val-Pro-Asn-Thr-Pro-Asn-Gln-Lys-Phe-Asp-Tyr-Glu-Val-Gly-Lys-Asp-Tyr-, and the molecular weight is 28,690. Intraperitoneal injection can induce a series of toxic symptoms of cockroaches and mouse; and a tetrodotoxin-resistant voltage-gated sodium ion channel on the nerve cell membrane can be inhibited. The invention also provides a purifying method of the toxin protein, which specifically comprises preparation of an egg extract, a molecular sieve chromatography step, an ion-exchange chromatography step and an inverse phase-high performance liquid chromatography step; and by introducing a C4 column with low hydrophobicity for purification, the steps are simple, and the purifying effect is good.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a toxin protein extracted from the ovum grains of the spider spider and a purification method thereof. Background technique [0002] L. tredecimguttatus, commonly known as black widow spider or black venomous spider, is a medium-sized spider that belongs to the phylum Arthropoda, Arachnida, Aranea, Ball Spideridae, and the genus Arachnida in taxonomy. . The venomous spider is one of the most poisonous spiders known in the world. Different from other poisonous animals, there is toxin not only in the venom of its venomous glands, but also in other tissues of the body (such as legs, abdominal tissues, etc.) and There are also toxic components in the eggs. It is of great theoretical and practical significance to study the toxic components outside the venom glands of A. [0003] Relevant foreign researchers have systematically and in-depth research on the venomous toxins of the spider ...

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Problems solved by technology

[0004] In 1971, Buffkin et al. used molecular sieve chromatography to divide the extract of the spider's egg into three main peaks, and proved that the first large peak component was toxic, but because it was not completely purified, it lacked specific information on the active components. Research

[0005] In 2001, Akhunov et al. also used molecular sieve chromatography to divide the egg grain extract of the spider spider into several parts, and proved that three of them were toxic, but they used ion exchange chromatography to further analyze the toxic part of the samples. When isolated, no toxic purified product was obtained, therefore, the research on the oogranulotoxin of the interspersed spider remained at the level of the crude extract

However, in actual work, RP-HPLC is generally not used for protein separation, because the hydrophobic interaction between proteins, especially those with large molecular weight, and the separation medium is strong, which may cause irreversible adsorption or denaturation of proteins.

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