Single-Molecule Imaging and Computational Microscopy Approaches Clarify the Mechanism of the Dimerization and Membrane Interactions of Green Fluorescent Protein

doi:10.3390/ijms20061410

	Single-Molecule Imaging and Computational Microscopy Approaches Clarify the Mechanism of the Dimerization and Membrane Interactions of Green Fluorescent Protein
	Wang, Xiaohua ; Song, Kai 1; Li, Yang 2; Tang, Ling 2; Deng, Xin
	2019
发表期刊	INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
ISSN	1661-6596
卷号	20 期号:6
摘要	Green fluorescent protein (GFP) is widely used as a biomarker in living systems; however, GFP and its variants are prone to forming low-affinity dimers under physiological conditions. This undesirable tendency is exacerbated when fluorescent proteins (FP) are confined to membranes, fused to naturally-oligomeric proteins, or expressed at high levels in cells. Oligomerization of FPs introduces artifacts into the measurement of subunit stoichiometry, as well as interactions between proteins fused to FPs. Introduction of a single mutation, A206K, has been shown to disrupt hydrophobic interactions in the region responsible for GFP dimerization, thereby contributing to its monomerization. Nevertheless, a detailed understanding of how this single amino acid-dependent inhibition of dimerization in GFP occurs at the atomic level is still lacking. Single-molecule experiments combined with computational microscopy (atomistic molecular dynamics) revealed that the amino group of A206 contributes to GFP dimer formation via a multivalent electrostatic interaction. We further showed that myristoyl modification is an efficient mechanism to promote membrane attachment of GFP. Molecular dynamics-based site-directed mutagenesis has been used to identify the key functional residues in FPs. The data presented here have been utilized as a monomeric control in downstream single-molecule studies, facilitating more accurate stoichiometry quantification of functional protein complexes in living cells.
关键词	single molecule stoichiometry molecular dynamics N-myristoylation
学科领域	Biochemistry & Molecular Biology ; Chemistry, Multidisciplinary
DOI	10.3390/ijms20061410
收录类别	SCI
语种	英语
WOS关键词	HIGH-THROUGHPUT ; FREE-ENERGY ; DYNAMICS ; SIMULATION ; OLIGOMERIZATION ; TRANSITION ; DEPENDENCE
WOS研究方向	Biochemistry & Molecular Biology ; Chemistry
WOS记录号	WOS:000465523400050
出版者	MDPI
文献子类	Article
出版地	BASEL
EISSN	1422-0067
资助机构	National Natural Science Foundation of ChinaNational Natural Science Foundation of China (NSFC) [31770400, 31270412] ; Chinese Academy of SciencesChinese Academy of Sciences
作者邮箱	wangxh@ibcas.ac.cn ; songkai@chd.edu.cn ; liyang184@mails.ucas.edu.cn ; tangling181@mails.ucas.edu.cn ; deng@ibcas.ac.cn
作品OA属性	Green Submitted, gold, Green Published
引用统计	被引频次：8[WOS] [WOS记录] [WOS相关记录]
文献类型	期刊论文
条目标识符	http://ir.ibcas.ac.cn/handle/2S10CLM1/19818
专题	中科院北方资源植物重点实验室
作者单位	1.Chinese Acad Sci, Inst Bot, Key Lab Plant Resources, Beijing 100093, Peoples R China 2.Changan Univ, Middle Sect South Er Huan Rd, Xian 710064, Shaanxi, Peoples R China 3.Univ Chinese Acad Sci, Beijing 100049, Peoples R China
推荐引用方式 GB/T 7714	Wang, Xiaohua,Song, Kai,Li, Yang,et al. Single-Molecule Imaging and Computational Microscopy Approaches Clarify the Mechanism of the Dimerization and Membrane Interactions of Green Fluorescent Protein[J]. INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES,2019,20(6).
APA	Wang, Xiaohua,Song, Kai,Li, Yang,Tang, Ling,&Deng, Xin.(2019).Single-Molecule Imaging and Computational Microscopy Approaches Clarify the Mechanism of the Dimerization and Membrane Interactions of Green Fluorescent Protein.INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES,20(6).
MLA	Wang, Xiaohua,et al."Single-Molecule Imaging and Computational Microscopy Approaches Clarify the Mechanism of the Dimerization and Membrane Interactions of Green Fluorescent Protein".INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES 20.6(2019).

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