用于抗蛇毒血清疗法的针对眼镜蛇毒液毒素的植物生产型 VHH 抗体的特征描述

Q1 Immunology and Microbiology

Biotechnology Reports Pub Date : 2024-04-16 DOI:10.1016/j.btre.2024.e00841

Sarocha Vitayathikornnasak , Kaewta Rattanapisit , Ashwini Malla , Pipob Suwanchaikasem , Richard Strasser , Narach Khorattanakulchai , Kanokporn Pothisamutyothin , Wanatchaporn Arunmanee , Waranyoo Phoolcharoen

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引用次数: 0

摘要

眼镜蛇（Naja kaouthia）毒液含有多种毒素，包括α-神经毒素（αNTX）和磷脂酶 A2（PLA2），可导致神经变性、呼吸衰竭甚至死亡。从动物血清中提取的传统抗蛇毒血清面临许多挑战和限制。融合了 VHH 与人类 IgG Fc 区的纯重链抗体（HCAb）在组织穿透、抗原结合和延长半衰期方面具有优势。本研究在烟草叶中构建并瞬时表达了两种针对α-神经毒素（VHH-αNTX-FC）和磷脂酶A2（VHH-PLA2-FC）的VHH-FC。重组 HCAbs 经过长达六天的培养以优化表达水平，然后用亲和层析法纯化，并使用 LC/Q-TOF 质谱法（MS）进行表征。纯化蛋白质的序列覆盖率超过 92%，平均质量约为 82 kDa，具有高甘露糖 N-聚糖特征。抗原结合试验表明，与 VHH-PLA2-Fc 相比，VHH-αNTX-Fc 与粗毒液的结合能力更强。

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Characterization of plant produced VHH antibodies against cobra venom toxins for antivenom therapy

Cobra (Naja kaouthia) venom contains many toxins including α-neurotoxin (αNTX) and phospholipase A2 (PLA2), which can cause neurodegeneration, respiratory failure, and even death. The traditional antivenom derived from animal serum faces many challenges and limitations. Heavy-chain-only antibodies (HCAb), fusing V_HH with human IgG Fc region, offer advantages in tissue penetration, antigen binding, and extended half-life. This research involved the construction and transient expression of two types of V_HH-F_C which are specific to α-Neurotoxin (V_HH-αNTX-F_C) and Phospholipase A2 (V_HH-PLA2-F_C) in Nicotiana benthamiana leaves. The recombinant HCAbs were incubated for up to six days to optimize expression levels followed by purification by affinity chromatography and characterization using LC/Q-TOF mass spectrometry (MS). Purified proteins demonstrated over 92 % sequence coverage and an average mass of around 82 kDa with a high-mannose N-glycan profile. An antigen binding assay showed that the V_HH-αNTX-Fc has a greater ability to bind to crude venom than V_HH-PLA2-Fc.

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来源期刊

Biotechnology Reports Immunology and Microbiology-Applied Microbiology and Biotechnology

CiteScore

15.80

自引率

0.00%

发文量

审稿时长

55 days

期刊介绍： Biotechnology Reports covers all aspects of Biotechnology particularly those reports that are useful and informative and that will be of value to other researchers in related fields. Biotechnology Reports loves ground breaking science, but will also accept good science that can be of use to the biotechnology community. The journal maintains a high quality peer review where submissions are considered on the basis of scientific validity and technical quality. Acceptable paper types are research articles (short or full communications), methods, mini-reviews, and commentaries in the following areas: Healthcare and pharmaceutical biotechnology Agricultural and food biotechnology Environmental biotechnology Molecular biology, cell and tissue engineering and synthetic biology Industrial biotechnology, biofuels and bioenergy Nanobiotechnology Bioinformatics & systems biology New processes and products in biotechnology, bioprocess engineering.