胰岛素瘤相关蛋白1（INSM1）是一类转录调节蛋白，通过其C-端的锌指结构域（氨基酸250-510）来识别序列特异性的DNA分子.INSM1的C-端包含有5个串联的锌指结构域，然而这些结构域的结构及其如何识别DNA的分子机制目前仍不清楚.通过重组构建的质粒pET-32m-INSM1（424-497）表达的蛋白质（氨基酸424-497）包含了最后两个锌指结构域4和5，简称为ZF（4-5）.该文详细研究了蛋白质ZF（4-5）的诱导表达条件，得到了较高产率的纯化蛋白.核磁共振（NMR）谱和圆二色谱（CD）揭示了Zn2+对稳定锌指蛋白结构的必要性，以及C2H2-Zn2+结合的组氨酸呈现为δ-异构方式.
刘买利
,
岳霞丽
,
朱勤俊
,
杨运煌
,
王华谱
. 蛋白质INSM1中的锌指结构域ZF (4-5)的表达、纯化与表征[J]. 波谱学杂志, 2017
, 34(1)
: 1
-7
.
DOI: 10.11938/cjmr20170101
Human insulinoma-associated protein 1 (INSM1) is a transcriptional regulator recognizing sequence-specific DNA through its C-terminal zinc finger (ZF) domains. INSM1 contains five zinc finger domains, whose structures are still not known; therefore, the mechanisms through which it recognizes DNA also remain unclear. In this study, we designed the recombinant plasmid pET-32m-INSM1(424-497), which can express the truncated INSM1 fragment containing the zinc finger domains 4 and 5[i.e., ZF(4-5)]. Expression and purification of ZF(4-5) were explored in order to achieve high protein yield for further structural and functional study. Nuclear magnetic resonance (NMR) and circular dichroism (CD) spectra revealed that chelation of Zn2+ to C2H2 in the ZF(4-5) was important for its structural stability, and also confirmed that the active-sites, Zn2+-chelated histidines, had the δ-tautomeric form.
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