澳大利亚棕伊澳蛇（Pseudechis australis）毒液中的磷脂酶A2——Pa-11的化学修饰对其生物学活性的影响。

Effects of chemical modifications of Pa-11, a phospholipase A2 from the venom of Australian king brown snake (Pseudechis australis), on its biological activities.

作者信息

Takasaki C, Sugama A, Yanagita A, Tamiya N, Rowan E G, Harvey A L

机构信息

Department of Chemistry, Faculty of Science, Tohoku University, Sendai, Japan.

出版信息

Toxicon. 1990;28(1):107-17. doi: 10.1016/0041-0101(90)90012-v.

DOI:10.1016/0041-0101(90)90012-v

PMID:2109907

Abstract

Pa-11, a phospholipase A2 isolated from the venom of an Australian elapid snake Pseudechis australis, was chemically modified and its enzymic, neuromuscular and lethal activities were studied. Carboxymethylation of Met-8 gave a derivative with 2% of the enzymic activity and less than 3% of the lethal activity of native Pa-11; it had about 5% of the original ability to block directly and indirectly stimulated mouse phrenic nerve-hemidiaphragm preparations. Nitrophenylsulfenylation of tryptophanyl residues at positions 31 and 69 caused loss of all activities. Amidination of all 14 lysyl residues gave a derivative with 41% and 16% of the enzymic and lethal activities, respectively, but with less than 5% of the original neuromuscular blocking activity. Mono-carbamoylation of lysyl residues at positions 58, 63, 81 and 85 was achieved. The most abundant derivative, 58-carbamoyl-lysine Pa-11 was enzymically 130% and lethally 100% as active as native Pa-11, but it had only about 20% of the native's neuromuscular activity in vitro. 63-Carbamoyl-lysine Pa-11 had 10% of the enzymic and 20% of the lethal activities, respectively; however, it retained at least 50% of its ability to block neuromuscular transmission in vitro, while losing most of its activity to block directly stimulated muscle contractions. 81- and 85-Carbamoyl derivatives have the same enzymic and lethal activities as the original protein, but the 85 derivative had less than 10% of the native neuromuscular activity. Hence, modifications of lysine residues at positions 58, 63 and 85 seem to be particularly significant in altering the neuromuscular, but not enzymic, activity of Pa-11, perhaps by altering the ability of the toxin to bind to its target on nerve and muscle membranes. Modification at position 63 appeared to lead to a dissociation of effects on neuromuscular transmission and directly on muscle cells.

摘要

从澳大利亚眼镜蛇（澳洲拟眼镜蛇）毒液中分离出的磷脂酶A2（Pa - 11）经过化学修饰，并对其酶活性、神经肌肉活性和致死活性进行了研究。对甲硫氨酸8进行羧甲基化得到一种衍生物，其酶活性为天然Pa - 11的2%，致死活性不到3%；它对直接和间接刺激的小鼠膈神经 - 半膈肌标本的阻断能力约为原来的5%。对第31位和第69位色氨酸残基进行硝基苯硫基化导致所有活性丧失。对所有14个赖氨酸残基进行脒基化得到一种衍生物，其酶活性和致死活性分别为原来的41%和16%，但神经肌肉阻断活性不到原来的5%。实现了对第58、63、81和85位赖氨酸残基的单氨甲酰化。最丰富的衍生物58 - 氨甲酰赖氨酸Pa - 11的酶活性是天然Pa - 11的130%，致死活性是100%，但在体外其神经肌肉活性仅为天然的约20%。63 - 氨甲酰赖氨酸Pa - 11的酶活性和致死活性分别为10%；然而，它在体外至少保留了50%的阻断神经肌肉传递的能力，同时大部分直接刺激肌肉收缩的活性丧失。81 - 和85 - 氨甲酰衍生物的酶活性和致死活性与原始蛋白相同，但85衍生物的神经肌肉活性不到天然的10%。因此，第58、63和85位赖氨酸残基的修饰似乎在改变Pa - 11的神经肌肉活性而非酶活性方面特别重要，这可能是通过改变毒素与神经和肌肉膜上靶点结合的能力实现的。第63位的修饰似乎导致了对神经肌肉传递和直接对肌肉细胞的作用的解离。