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首页> 外文期刊>Journal of bacteriology >Assembly of the aspartate transcarbamoylase holoenzyme from transcriptionally independent catalytic and regulatory cistrons.
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Assembly of the aspartate transcarbamoylase holoenzyme from transcriptionally independent catalytic and regulatory cistrons.

机译：天冬氨酸转氨甲酰酶全酶由转录独立的催化和调节顺反子组装而成。

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摘要

The cistrons encoding the regulatory and catalytic polypeptides of aspartate transcarbamoylase (EC 2.1.3.2) from Escherichia coli K-12 have been cloned separately on plasmids from different incompatability groups. The catalytic cistron (pyrB) was carried by pACYC184 and expressed from its own promoter, whereas the regulatory cistron was expressed from the lac po of pBH20. The catalytic polypeptide chains assembled into enzymatically active trimers (c3) in vivo when expressed in the absence of regulatory subunits. Similarly, the regulatory polypeptide chains assembled into regulatory dimers (r2) in vivo in the absence of catalytic subunits. When cellular extracts containing regulatory dimers and catalytic trimers synthesized in separate cells were combined in vitro, partial spontaneous holoenzyme assembly occurred. When pyrB and pyrI were expressed from transcriptionally independent cistrons in the same cell, all detectable catalytic polypeptides were incorporated into the functional aspartate transcarbamoylase holoenzyme, 2(c3):3(r2). Thus, it is clear that the in vivo assembly of ATCase holoenzyme is a direct, spontaneous process involving the association of preformed regulatory subunits (r2) and catalytic subunits (c3). This procedure provides a general method for the construction of hybrid aspartate transcarbamoylase in vivo and may be applicable to other oligomeric enzymes constructed from different polypeptides.
机译：已经将编码来自大肠杆菌K-12的天冬氨酸转氨甲酰酶的调节性和催化性多肽（EC 2.1.3.2）的顺反子分别克隆到来自不同不相容性组的质粒上。催化顺反子（pyrB）由pACYC184携带并由其自身的启动子表达，而调节性顺反子则由pBH20的紫胶表达。当在不存在调节亚基的情况下表达时，催化多肽链在体内组装成酶活性三聚体（c3）。类似地，在不存在催化亚基的情况下，调节多肽链在体内组装成调节二聚体（r2）。当在体外将包含在单独细胞中合成的调节二聚体和催化三聚体的细胞提取物合并时，会发生部分自发的全酶组装。当在同一细胞中从转录独立的顺反子表达pyrB和pyrI时，所有可检测的催化多肽都被掺入功能性天冬氨酸转氨甲酰酶全酶2（c3）：3（r2）中。因此，很明显，ATCase全酶的体内组装是直接的，自发的过程，涉及预先形成的调节亚基（r2）和催化亚基（c3）的缔合。该程序提供了在体内构建杂合天冬氨酸转氨甲酰酶的一般方法，并且可适用于由不同多肽构建的其他寡聚酶。

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