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功能活跃的人类SLC38A2 (SNAT2) 的细菌过度生产利用神秘标签:一种便宜,快速的工具来测试连接体

Michele Galluccio¹, Martina Tripicchio¹, Lara Console¹

¹Department DiBEST (Biologia, Ecologia, Scienze Della Terra) Laboratory of Biochemistry, Molecular Biotechnology and Molecular Biology, University of Calabria, Via P. Bucci 4C-6C, 87036, Arcavacata di Rende, Italy.

Molecular biology reports

|February 23, 2024

概括

研究人员使用Mistic标签成功过度表达并净化了SLC38A2氨基酸载体. 这种纯化的功能转运体对于未来的结构-功能研究和连接体识别至关重要.

科学领域:

生物化学生物化学
分子生物学分子生物学
膜运输运输膜运输

背景情况:

SLC38A2是中性氨基酸的Na+依赖的载体.
它在II型糖尿病和癌症中起作用.
关于其结构-功能关系和监管连接物的知识有限.

研究的目的:

为了过度表达和净化SLC38A2传送器.
为了获得功能性蛋白质进行进一步的研究.
为了使结构-功能关系的研究和连接体查.

主要方法:

将SLC38A2cDNA克隆成一个pET-28-Mistic向量.
BL21编码子加上RIL大肠杆菌菌株的转化.
用葡萄糖和氧气优化表达;通过-化亲和力染色法进行净化.
将纯化的hSNAT2溶解成蛋白质体.

主要成果:

成功过度表达和净化hSNAT2-Mistic仿真体,产量为~60 mg/L.
神秘标签对于高效的表达和净化至关重要.
纯化的hSNAT2被重新构成蛋白质体,以活性,右侧向外的方向.

结论:

N端的Mistic标签促进了hSNAT2的过度表达和净化.
纯化的,功能性的hSNAT2传送器是一个有价值的工具.
允许未来的结构功能研究和新兴抑制剂/激活剂的识别.

关键词:

蛋白质体蛋白质体净化净化净化净化再折叠可以重新折叠.在SLC中,SLC是SLC.在SNAT2中,SNAT2是SNAT2.

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