酵母KEX2内酶正确地在哺乳动物细胞中切割神经内分泌前激素
G Thomas1, B A Thorne, L Thomas
1Vollum Institute for Advanced Biomedical Research, Oregon Health Sciences University, Portland 97201.
概括
缺乏特定酶的哺乳动物细胞系无法处理前体激素的前体. 引入酵母KEX2基因使得对小鼠亲米兰皮质蛋白 (mPOMC) 的处理成为可能,证明了酶的存在.
科学领域:
- 分子生物学分子生物学
- 细胞生物学 细胞生物学
- 生物化学 生物化学
背景情况:
- 哺乳动物细胞系往往缺乏用于前激素前体加工所需的内源性酶.
- 鼠类神经内分泌的前体,前-阿片梅拉诺科丁 (mPOMC),需要特定的内酶来成熟.
- 了解前激素处理对于研究激素调节和功能至关重要.
研究的目的:
- 调查特定内酶在前体激素前体加工中的作用.
- 为了确定是否引入异质加工酶可以在缺陷细胞系中挽救mPOMC成熟.
- 为了确定细胞特异性前激素加工的酶基础.
主要方法:
- 在哺乳动物细胞系中利用复合疫苗病毒载体进行基因表达 (BSC-40,NG108-15,GH4C1).
- 与疫苗病毒载体共感染的细胞编码了酵母KEX2基因,这是已知的内酶.
- 使用生物化学方法分析了mPOMC的细胞内分裂产物.
主要成果:
- 与表达KEX2的病毒的同时感染使得以前缺乏的细胞系中mPOMC的细胞内分裂成为可能.
- 产生了成熟产品,包括-利波和β-endorphin1-31.
- 在GH4C1细胞中,加工的被正确地分类成分泌颗粒.
结论:
- 细胞系无法处理益激素的原因很可能是由于缺乏特定的内源性加工酶.
- 酵母KEX2内酶的异质表达可以恢复前激素处理.
- 这突显了特定蛋白酶在前体的翻译后修改中的关键作用.
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