DNA G-四链体的结构完整性对催化性质的影响

doi:10.1016/S1872-2067(20)63744-5

催化学报 ›› 2021, Vol. 42 ›› Issue (7): 1102-1107.DOI: 10.1016/S1872-2067(20)63744-5

DNA G-四链体的结构完整性对催化性质的影响

陈杰林^a,^†, 程明攀^a,^†, 王佳伟^a, 仇得辉^a, David Monchaud^b, Jean-Louis Mergny^a^,^c, 鞠熀先^a^,^*(), 周俊^a^,^#()

^a南京大学化学化工学院, 生命分析化学国家重点实验室, 江苏南京210023, 中国
^b勃艮第大学分子化学研究所, 法国
^c巴黎综合理工学院, 光学与生物科学实验室, 法国

收稿日期:2020-10-10 接受日期:2020-11-11 出版日期:2021-07-18 发布日期:2020-12-10
通讯作者: 鞠熀先,周俊
作者简介:
^†共同第一作者.
基金资助:
国家自然科学基金(21977045);国家自然科学基金(21635005);中央高校基本科研业务费(02051430210);南京大学高层次人才科研启动经费(020514912216)

The catalytic properties of DNA G-quadruplexes rely on their structural integrity

Jielin Chen^a,^†, Mingpan Cheng^a,^†, Jiawei Wang^a, Dehui Qiu^a, David Monchaud^b, Jean-Louis Mergny^a^,^c, Huangxian Ju^a^,^*(), Jun Zhou^a^,^#()

^aState Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210023, Jiangsu, China
^bInstitut de Chimie Moléculaire, Université de Bourgogne (ICMUB), CNRS UMR6302, UBFC Dijon 21000, France
^cLaboratoire d’Optique et Biosciences, Ecole Polytechnique, CNRS, INSERM, Institut Polytechnique de Paris, 91128 Palaiseau, France

Received:2020-10-10 Accepted:2020-11-11 Online:2021-07-18 Published:2020-12-10
Contact: Huangxian Ju,Jun Zhou
About author:^# E-mail: jun.zhou@nju.edu.cn
^* E-mail: hxju@nju.edu.cn;
First author contact:
^† These authors contributed equally to this work.
Supported by:
National Natural Science Foundation of China(21977045);National Natural Science Foundation of China(21635005);Fundamental Research Funds for the Central Universities(02051430210);Funds of Nanjing University(020514912216)

摘要/Abstract

摘要：

DNA酶中的G-四链体-血红素(G4-hemin)DNA酶结构具有较高的设计性和化学稳定性, 因此格外受研究者关注. G-平面作为辅酶因子hemin的结合位点, 不仅提供大π平面与hemin结合, 而且其平面上的G碱基还可以充当近端配位基团与hemin进行配位. 因此, 研究G-平面完整性在G4-DNA酶体系中的作用具有重要意义. 本文设计了一系列含有空位的G4(G-vacancy, GV)及G-三链体, 通过“鸟嘌呤类似物插入”策略实现G-平面完整性以及DNA酶催化活性的恢复. 结果表明, 末端G-平面完整性是G4-DNA酶具有催化活性的必要条件, 且其能够充当近端配位基团与末端碱基协同激活G4-DNA酶.
考虑到hemin会选择性地结合于G4的3’-端平面, 本文以含有3’-端空位的G4以及G-三链体为模型进行DNA酶的构建. 结果表明,相较于末端完整的G4-hemin DNA酶, 末端不完整的G4结构所形成的DNA酶催化活性很低. 为了进一步验证该平面完整性的重要性, 本文提出了“鸟嘌呤衍生物插入”策略, 即将鸟嘌呤衍生物(无环鸟苷和鸟苷)插入G-空位以恢复G-平面的完整性. 通过圆二色光谱和紫外熔解实验, 发现末端平面完整性的缺失会使圆二色特征峰信号和G4结构热稳定性下降, 而鸟嘌呤碱基类似物的加入则可以使特征峰信号以及热稳定性得到一定程度的恢复, 表明鸟嘌呤碱基类似物的加入确实使G-平面完整性得到恢复. 与此同时, 随着鸟嘌呤碱基类似物浓度的增加, G4-hemin DNA酶活性逐渐增强, 最终恢复至与完整G4一样的活性. 在以G-三链体为模型的实验中, 本文通过另一条富G序列与G-三链体进行结合, 形成复合的(3+1)型G4结构, 最终实现了DNA酶活性的恢复. 同时, 在3’-G-平面末端增加了激活碱基(dA或dTC), 结果表明, 即使G-平面不完整, 末端碱基依旧能够激活DNA酶, 但酶活性整体弱于完整G4时的活性. 同样, “鸟嘌呤衍生物插入”策略可以使酶活性得到恢复.
本文系列实验充分说明了末端碱基可与G-平面形成协同作用, 与hemin的铁中心共同形成六配位关系, 加速催化中间体生成, 进而增强催化活性. 有趣的是, 通过设计Holliday junction结构研究发现, “鸟嘌呤衍生物插入”策略仅适用于平行G4结构. G-空位的存在不仅降低了G4结构的稳定性, 而且降低了其与hemin间的亲和力, 二者均是造成G4-DNA酶催化能力下降的主要因素. 总之, 本文证明了3’-端G-平面的完整性是G4-DNA酶实现其催化能力必不可少的因素, 对理解末端G-平面在G4-DNA酶中的作用具有重要的参考意义.

关键词: G-四链体, G-四链体DNA酶, G-空位, G-平面完整性, 鸟嘌呤衍生物

Abstract:

The influence of the G-quartet structural integrity on the catalytic activity of the G-quadruplex (G4) was investigated by comparing the G4-DNAzyme performances of a series of G4s with a G-vacancy site and a G-triplex (G-tri). The results presented herein not only confirm that the structural integrity of the 3'-end G-quartet is necessary for G4s to be catalytically competent but also show how to remediate G-vacancy-mediated catalytic activity losses via the addition of guanine surrogates in an approach referred to as G-vacancy complementation strategy that is applicable to parallel G4s only. Furthermore, this study demonstrates that the terminal G-quartet could act as a proximal coordinating group and cooperate with the flanking nucleotide to activate the hemin cofactor.

Key words: G-quadruplex, G-quadruplex DNAzyme, G-vacancy, G-quartet integrity, Guanine surrogate

陈杰林, 程明攀, 王佳伟, 仇得辉, David Monchaud, Jean-Louis Mergny, 鞠熀先, 周俊. DNA G-四链体的结构完整性对催化性质的影响[J]. 催化学报, 2021, 42(7): 1102-1107.

Jielin Chen, Mingpan Cheng, Jiawei Wang, Dehui Qiu, David Monchaud, Jean-Louis Mergny, Huangxian Ju, Jun Zhou. The catalytic properties of DNA G-quadruplexes rely on their structural integrity[J]. Chinese Journal of Catalysis, 2021, 42(7): 1102-1107.

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图/表 6

Table 1 DNA sequences used in this study.

Name	Sequences (5′-3′)
GV1	TT GTG T GGG T GGG T GGG T
GV2	TT GG T GGG T GGG T GGG T
GV2A	TT GG T GGG T GGG T GGG A
GV2TC	TT GG T GGG T GGG T GGG TC
PG4	TT GGG T GGG T GGG T GGG T
G4A	TT GGG T GGG T GGG T GGG A
G4TC	TT GGG T GGG T GGG T GGG TC
G-Tri	TT GGG T GGG T GGG
TG3T	T GGG T

Fig. 1. Schematic representation of the influence of the G-quartet integrity on the G4-DNAzyme activity. Upper panel: G4 with a G-vacancy site (GV) (structure of GV2 is shown here, PDB ID is 6K3X) can be completed by the free guanine surrogates [(acyclovir (A) and guanosine (G)]. Lower panel: the G-quartet integrity of a G-triplex (G-Tri) was restored with the single-stranded G-rich sequence TG3T.

Fig. 2. (a) G-vacancy complementation strategy with free guanine surrogates. G4-DNAzyme activity of (b) GVs and G4s, and (c) GV1, (d) GV2, (e) GV2A, and (f) GV2TC upon addition of increasing concentrations (2, 5, 20, 50, and 100 mol. equiv.) of guanine surrogates [acyclovir (A) and guanosine (G)].

Fig. 3. G4-DNAzyme activity of G-triplex (G-Tri) upon addition of increasing concentrations (5, 10, 20, and 50 mol equiv.) of TG3T.

Fig. 4. Catalytic performances of Holliday junction-constrained G4s: (a) Schematic representation of the seven different Holliday junction-induced G4 systems (HGV and HG4): with (HG4, HG4TC-1, and HG4TC-2) or without (HGV1, HGV1TC, HGV2, and HGV2TC) intact G-quartet, and some of them with flanking dTC (HGV1TC, HGV2TC, HG4TC-1, and HG4TC-2). (b) The G4-DNAzyme activity of GVs upon addition of increasing concentrations of acyclovir (A).

Fig. 5. Dissociation constants (Kd) between hemin and GVs (red bars), GVs upon addition of increasing concentrations of acyclovir (A) (green bars), and intact G4s (blue bars). Student’s t-tests were performed between intact G4s and GVs (containing GV2 integrated with acyclovir): *** means p < 0.001, i.e. the difference between each group data set is very significant.

参考文献

[1]	R. R. Breaker, G. F. Joyce, Chem. Biol., 1994,1, 223-229.
[2]	Z. Li, C. Wang, J. Li, J. Zhang, C. Fan, I. Willner, H. Tian, CCS Chem., 2020,2, 707-728.
[3]	L. Liu, J. Lin, Y. Song, C. Yang, Z. Zhu, Chem. Res. Chin. Univ., 2020,36, 247-253.
[4]	S. K. Silverman, Trends Biochem. Sci., 2016,41, 595-609.
[5]	A. J. Boersma, R. P. Megens, B. L. Feringa, G. Roelfes, Chem. Soc. Rev., 2010,39, 2083-2092.
[6]	M. Cheng, Y. Li, J. Zhou, G. Jia, S. M. Lu, Y. Yang, C. Li, Chem. Commun., 2016,52, 9644-9647.
[7]	I. Willner, B. Shlyahovsky, M. Zayats, B. Willner, Chem. Soc. Rev., 2008,37, 1153-1165.
[8]	J. Liu, Z. Cao, Y. Lu, Chem. Rev., 2009,109, 1948-1998.
[9]	L. Ma, J. Liu, iScience, 2020,23, 100815.
[10]	J. H. Yum, S. Park, H. Sugiyama, Org. Biomol. Chem., 2019,17, 9547-9561.
[11]	J. -L. Mergny, D. Sen, Chem. Rev., 2019,119, 6290-6325.
[12]	P. Travascio, Y. Li, D. Sen, Chem. Biol., 1998,5, 505-517.
[13]	S. Nakayama, H. O. Sintim, Anal. Chim. Acta, 2012,747, 1-6.
[14]	L. Stefan, F. Denat, D. Monchaud, Nucleic Acids Res., 2012,40, 8759-8772.
[15]	D. Sen, L. C. H Poon, Crit. Rev. Biochem. Mol. Biol., 2011,46, 478-492.
[16]	L. Stefan, F. Denat, D. Monchaud, J. Am. Chem. Soc., 2011,133, 20405-20415.
[17]	Q. Liu, H. Wang, X. Shi, Z. G. Wang, B. Ding, ACS Nano, 2017,11, 7251-7258.
[18]	W. Li, Y. Li, Z. Liu, B. Lin, H. Yi, F. Xu, Z. Nie, S. Yao, Nucleic Acids Res., 2016,44, 7373-7384.
[19]	J. Chen, Y. Guo, J. Zhou, H. Ju, Chem. Eur. J., 2017,23, 4210-4215.
[20]	T. Chang, H. Gong, P. Ding, X. Liu, W. Li, T. Bing, Z. Cao, D. Shangguan, Chem. Eur. J., 2016,22, 4015-4021.
[21]	J. Chen, Y. Zhang, M. Cheng, Y. Guo, J. Šponer, D. Monchaud, J.-L. Mergny, H. Ju, J. Zhou, ACS Catal., 2018,8, 11352-11361.
[22]	W. Li, S. Chen, D. Xu, Q. Wen, T. Yang, J. Liu, Chem. Eur. J., 2018,24, 14500-14505.
[23]	M. Cheng, J. Zhou, G. Jia, X. Ai, J.-L. Mergny, C. Li, Biochim. Biophys. Acta, Gen. Subj., 2017,1861, 1913-1920.
[24]	E. Golub, H. B. Albada, W. C. Liao, Y. Biniuri, I. Willner, J. Am. Chem. Soc., 2016,138, 164-172.
[25]	J. Chen, J. Wang, S. C C. van der Lubbe, M. Cheng, D. Qiu, D. Monchaud, J.-L. Mergny, C. F. Guerra, H. Ju, J. Zhou, CCS Chem., 2020,2, 2183-2193.
[26]	N. Shumayrikh, Y. C. Huang, D. Sen, Nucleic Acids Res., 2015,43, 4191-4201.
[27]	P. Travascio, P. K. Witting, A. G. Mauk, D. Sen, J. Am. Chem. Soc., 2001,123, 1337-1348.
[28]	R. Shinomiya, Y. Katahira, H. Araki, T. Shibata, A. Momotake, S. Yanagisawa, T. Ogura, A. Suzuki, S. Neya, Y. Yamamoto, Biochemistry, 2018,57, 5930-5937.
[29]	Y. Yamamoto, H. Araki, R. Shinomiya, K. Hayasaka, Y. Nakayama, K. Ochi, T. Shibata, A. Momotake, T. Ohyama, M. Hagihara, H. Hemmi, Biochemistry, 2018,57, 5938-5948.
[30]	I. Ourliac-Garnier, M. A Elizondo-Riojas, S. Redon, N. P. Farrell, S. Bombard, Biochemistry, 2005,44, 10620-10634.
[31]	H. Bertrand, S. Bombard, D. Monchaud, M. P Teulade-Fichou, J. Biol. Inorg. Chem., 2007,12, 1003-1014.
[32]	P. Stadlbauer, M. Krepl, T. E. Cheatham, 3rd, J. Koca, J. Šponer, Nucleic Acids Res., 2013,41, 7128-7143.
[33]	J. Wang, M. Cheng, J. Chen, H. Ju, D. Monchaud, J.-L. Mergny, J. Zhou, Chem. Commun., 2020,56, 1839-1842.
[34]	B. Heddi, N. Martin-Pintado, Z. Serimbetov, T. M. A Kari, A. T. Phan, Nucleic Acids Res., 2016,44, 910-916.
[35]	K. B. Wang, J. Dickerhoff, G. Wu, D. Yang, J. Am. Chem. Soc., 2020,142, 5204-5211.
[36]	F. R. Winnerdy, P. Das, B. Heddi, A. T. Phan, J. Am. Chem. Soc., 2019,141, 18038-18047.
[37]	Y. Guo, J. Chen, M. Cheng, D. Monchaud, J. Zhou, H. Ju, Angew. Chem. Int. Ed., 2017,56, 16636-16640.
[38]	X. M. Li, K. W. Zheng, Y. H. Hao, Z. Tan, Angew. Chem. Int. Ed., 2016,55, 13759-13764.
[39]	Y. D. He, K. W. Zheng, C. J. Wen, X. M. Li, J. Y. Gong, Y. H. Hao, Y. Zhao, Z. Tan, J. Am. Chem. Soc., 2020,142, 11394-11403.
[40]	D. J. Y Tan, P. Das, F. R. Winnerdy, K. W. Lim, A. T. Phan, Chem. Commun., 2020,56, 5897-5900.
[41]	S. Wang, B. Fu, S. Peng, X. Zhang, T. Tian, X. Zhou, Chem. Commun., 2013,49, 7920-7922.
[42]	N. Zhang, A. T. Phan, D. J. Patel, J. Am. Chem. Soc., 2005,127, 17277-17285.
[43]	O. Lustgarten, R. Carmieli, L. Motiei, D. Margulies, Angew. Chem. Int. Ed., 2019,58, 184-188.

DNA G-四链体的结构完整性对催化性质的影响

The catalytic properties of DNA G-quadruplexes rely on their structural integrity

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