Recombinant Butelase-Mediated Cyclization of the p53-Binding Domain of the Oncoprotein MdmX-Stabilized Protein Conformation as a Promising Model for Structural Investigation

doi:10.1021/acs.biochem.9b00263

	Recombinant Butelase-Mediated Cyclization of the p53-Binding Domain of the Oncoprotein MdmX-Stabilized Protein Conformation as a Promising Model for Structural Investigation
	Pi, Ni 1,4; Gao, Meng 1,4; Cheng, Xiyao 1,3,4; Liu, Huili 2; Kuang, Zhengkun 1,4; Yang, Zixin 1,4; Yang, Jing 1,4; Zhang, Bailing 1,4; Chen, Yao 1,4; Liu, Sen 1,4
刊名	BIOCHEMISTRY
	2019-07-09
卷号	58 期号:27 页码:3005-3015
ISSN号	0006-2960
DOI	10.1021/acs.biochem.9b00263
英文摘要	Cyclization of the polypeptide backbone has proven to be a powerful strategy for enhancing protein stability for fundamental research and pharmaceutical application. The use of such an approach is restricted by how well a targeted polypeptide can be efficiently ligated. Recently, an Asx-specific peptide ligase identified from a tropical cyclotide-producing plant and named butelase 1 exhibited excellent cyclization kinetics that cannot be matched by other known ligases, including intein, PATG, PCY1, and sortase A. In this work, we aimed to examine whether butelase 1 facilitated conformational stability for structural investigation First we successfullly expressed recombined butelase 1 (rBTase) in the yeast Pichia pastoris. Next, rBTase was shown to be highly efficient in the cyclization of the p53-binding domain (N-terminal domain) of murine double minute X (N-MdmX), an important target for designing anticancer drugs. The cyclized N-MdmX (cMdmX) exhibited increased conformational stability and improved interaction with the ligand compared with those of noncyclized N-MdmX. Importantly, the thermal melting process was completely reversible, contrary to noncyclized N-MdmX, and the melting temperature (T-m) of cMdmX was increased to 47 from 43 degrees C. This stable conformation of cMdmX was further confirmed by N-15-H-1 heteronuclear single-quantum coherence nuclear magnetic resonance (NMR) spectroscopy. The complex of cMdmX and the ligand was tested for protein crystallization, and several promising findings were revealed. Therefore, our work not only provides a recombinant version of butelase 1 but also suggests a conventional approach for preparing stable protein samples for both protein crystallization and NMR structural investigation.
资助项目	National Natural Science Foundation of China[21603121] ; National Natural Science Foundation of China[3167076831500132] ; State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics[T152604] ; Wuhan Science and Technology Bureau of China[2018060401011319]
WOS关键词	PEPTIDES ; MACROCYCLIZATION ; INHIBITOR ; LIGATION ; BINDING ; P53
WOS研究方向	Biochemistry & Molecular Biology
语种	英语
出版者	AMER CHEMICAL SOC
WOS记录号	WOS:000475408700006
资助机构	National Natural Science Foundation of China ; National Natural Science Foundation of China ; State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics ; State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics ; Wuhan Science and Technology Bureau of China ; Wuhan Science and Technology Bureau of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics ; State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics ; Wuhan Science and Technology Bureau of China ; Wuhan Science and Technology Bureau of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics ; State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics ; Wuhan Science and Technology Bureau of China ; Wuhan Science and Technology Bureau of China ; National Natural Science Foundation of China ; National Natural Science Foundation of China ; State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics ; State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics ; Wuhan Science and Technology Bureau of China ; Wuhan Science and Technology Bureau of China
内容类型	期刊论文
源URL	[http://ir.wipm.ac.cn/handle/112942/14643]
专题	中国科学院武汉物理与数学研究所
通讯作者	Huang, Yongqi; Su, Zhengding
作者单位	1.Hubei Univ Technol, Hubei Key Lab Ind Microbiol, Minist Educ, Key Lab Ind Fermentat, Wuhan 430068, Hubei, Peoples R China 2.Chinese Acad Sci, Wuhan Inst Phys & Math, State Key Lab Magnet Resonance & Atom & Mol Phys, Natl Ctr Magnet Resonance Wuhan, Wuhan 430071, Hubei, Peoples R China 3.Wuhan Amersino Biodevelop Inc, B1 Bldg,Biolake Pk, Wuhan 430075, Hubei, Peoples R China 4.Hubei Univ Technol, Natl Ctr Cellular Regulat & Mol Pharmaceut 111, Wuhan 430068, Hubei, Peoples R China
推荐引用方式 GB/T 7714	Pi, Ni,Gao, Meng,Cheng, Xiyao,et al. Recombinant Butelase-Mediated Cyclization of the p53-Binding Domain of the Oncoprotein MdmX-Stabilized Protein Conformation as a Promising Model for Structural Investigation[J]. BIOCHEMISTRY,2019,58(27):3005-3015.
APA	Pi, Ni.,Gao, Meng.,Cheng, Xiyao.,Liu, Huili.,Kuang, Zhengkun.,...&Su, Zhengding.(2019).Recombinant Butelase-Mediated Cyclization of the p53-Binding Domain of the Oncoprotein MdmX-Stabilized Protein Conformation as a Promising Model for Structural Investigation.BIOCHEMISTRY,58(27),3005-3015.
MLA	Pi, Ni,et al."Recombinant Butelase-Mediated Cyclization of the p53-Binding Domain of the Oncoprotein MdmX-Stabilized Protein Conformation as a Promising Model for Structural Investigation".BIOCHEMISTRY 58.27(2019):3005-3015.