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增强微生物转质氨酶的基质特异性,用于精确的纳米体标签

Xinglong Wang^1,2,3, Kangjie Xu³, Haoran Fu³

¹School of Food Science and Technology, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu, 214122, China.

Synthetic and systems biotechnology

|November 18, 2024

概括

工程化Streptomyces mobaraenesis transglutaminase (smTG) 能够准确地标记特定位置的蛋白质. 经过修改的smTG变体促进了用于先进的生物结合应用的纳米体-合物结合物的创建.

科学领域:

生物化学生物化学
分子生物学分子生物学
蛋白质工程是指蛋白质工程.

背景情况:

链杆菌的mobaraenesis转胺酶 (smTG) 用于蛋白质标签.
现有的smTG方法可以导致非特异性蛋白质交叉链接.
精确的特定位点标签对于诸如抗体-药物联体等应用至关重要.

研究的目的:

为了设计smTG,准确地对蛋白质进行特定位置的标记.
开发一种方法来制造纳米体-化物合物 (NFC).
为了增强smTG的活性,针对特定的基质进行向的合.

主要方法:

利用分子对接和虚拟突变发生来重新设计smTG基质特异性.
引入了单点突变 (G250H,Y278E) 进入一个热稳定的smTG变体 (TGm2).
对GGGGQR和标准基质进行了酶活性评估.
使用工程 smTG.使用光体对纳米体 (1C12,7D12) 进行现场特定标记.

主要成果:

突变G250H和Y278E显著增强了smTG对GGGQR的活性.
Y278E突变极大地改变了基质偏好,使GGGGQR活性比从0.05增加到0.93.
工程 smTG 变体 TGm2-Y278E 成功实现了与 GGGGQR 融合的纳米体的特定位点标记.
与其他smTG变体观察到非特异性标签,突出显示了TGm2-Y278E的特异性.

结论:

通过向突变工程 smTG 可以实现精确的特定位点蛋白质标记.
修改后的smTG变体是纳米体-相结合物的生物合成的有效工具.
这种方法对抗体-药物结合物和其他生物结合物的开发具有前景.

关键词:

检测过敏原检测检测过敏原检测酶 - 连合体对接对接纳米体与药物的结合.链杆菌的mobaraenesis转谷氨酸酶.虚拟突变发生是虚拟的突变发生.

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