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2019年12月,由新型冠状病毒(SARS-CoV-2)引起的新型冠状病毒肺炎(COVID-19)在中国武汉暴发。SARS-CoV-2的基因组编码2种病毒蛋白酶,即木瓜样蛋白酶(Papain-like protease,PLpro)和3C样蛋白酶(3C-like protease)。其中PLpro是SARS-CoV-2复制酶复合体(RC)形成的重要调节蛋白分子,对于病毒基因组转录和复制至关重要。因此,将SARS-CoV-2 PLpro作为药物的靶点对COVID-19的治疗具有积极意义。本研究应用生物信息学工具分析新型冠状病毒的木瓜样蛋白酶的结构和功能,首先利用BLAST和BioEdit获取SARS-CoV-2 PLpro蛋白酶(SC2-PLpro)及其同源蛋白的氨基酸序列,并利用BLAST和MEGA 6.0进行同源性分析。之后,利用ProParam和Proscale分别对SC2-PLpro蛋白酶的理化性质、亲水性和疏水性进行分析。然后,通过SOMPA、ScanProsite和InterPro分别预测SC2-PLpro蛋白酶的二级结构和功能区域,进一步利用SignalP 4.0和TMHMM对SC2-PLpro蛋白酶的信号肽和跨膜区进行分析。最后,通过SWISS-MODEL对SARS-CoV-2 PLpro蛋白酶进行三级结构同源建模。结果显示,对SARS-CoV-2 PLpro蛋白酶与已报道的PLpro蛋白酶进行多序列比对后,发现SARS-CoV-2 nsp3的746～1 063段氨基酸与多种冠状病毒PLpro蛋白酶氨基酸序列高度相似。同时,同源性分析发现SARS-CoV-2与蝙蝠冠状病毒的PLpro蛋白酶具有同源性,其中与QHR63299、AVP78030相似性最高。对SC2-PLpro进行理化性质预测结果显示,其由318个氨基酸所组成,为稳定亲水性蛋白。二级结构预测结果显示SC2-PLpro主要含有α-螺旋、延伸链、β-转角、无规卷曲,四种结构贯穿整条氨基酸链。进一步进行功能分析,发现其具有完整的催化三联体、锌结合域、泛素样N末端结构域,故推测该蛋白具有去泛素化的功能。然后,信号肽假说和跨膜结构域分析结果表明,SC2-PLpro既不是分泌蛋白,也不属于跨膜蛋白。本研究提示,生物信息学分析SC2-PLpro为稳定性亲水蛋白,属于非跨膜蛋白,比较保守,具有去泛素化的功能,利用此功能可以进一步规避宿主的固有免疫反应。通过制备PLpro蛋白酶小分子抑制剂,可能有助于治疗新型冠状病毒肺炎。
Abstract：Since December 2019,the coronavirus disease 2019(COVID-19)caused by SARS-CoV-2 broke out in Wuhan,China. The genome of SARS-CoV-2 encodes two viral proteases,including papain-like protease(PLpro)and 3 C-like protease(3 C-like protease). Among them,PLpro is a significant regulatory factor formed by SARS-CoV-2 replicase complex(RC),and it is essential for viral genome transcription and replication.Therefore,SARS-CoV-2 PLpro as an attractive drug target for treating COVID-19 has an important meaning.To predict the structure and function of PLpro of SARS-CoV-2(SC2-PLpro),the amino-acid sequences of SC2-PLpro and its homologous viruses were obtained by BLAST and BioEdit. The homology was analyzed by BLAST and MEGA 6.0. The basic physicochemical properties and hydrophilicity of SC2-PLpro were predicted by ProParam and Proscale. Then,the secondary structure and functional regions of SC2-PLpro were predicted by SOMPA,ScanProsite and InterPro,respectively. The SignalP 4.0 Server and TMHMM were employed to predict the signaling-peptide and transmembrane regions of SC2-PLpro. Finally,the tertiary structure of SC2-PLpro was constructed using SWISS-MODEL. Results showed that after multiple sequence alignments of SC2-PLpro and other PLpro proteases had been reported,it was found that 746 – 1063 amino acids of nsp3 from SARS-CoV-2 were highly similar to those of various PLpros. Homology analyses revealed that SC2-PLpro had homology with PLpros from most bat coronaviruses,and that it had the highest similarity with QHR63299 and AVP78030. Prediction of the physicochemical properties of SC2-PLpro showed that it was composed of 318 amino acids and a stable hydrophilic protein. Prediction of the secondary structure revealed that SC2-PLpro mainly contained an α-helix,extended chain,β-turn and random coil. Four structures penetrated the entire amino-acid chain. Functional analyses demonstrated that it had a complete catalytic triad,zinc-binding domain,and ubiquitin-like N-terminal domain,which suggested that it had the function of deubiquitination. Based on analyses of signaling peptides and transmembrane domains,SC2-PLpro is neither a secretory protein nor a transmembrane protein. In the present study,bioinformatics analysis showed that SC2-PLpro was a stable,hydrophilic and non-transmembrane protein. PLpro was relatively conserved,and this protease had the function of deubiquitination to avoid the innate immune response of the host. PLpro inhibitors might be helpful to treat COVID-19.
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基本信息:
DOI：10.13242/j.cnki.bingduxuebao.003841
中图分类号:R373
引用信息:
[1]杜倩倩,张智慧,廉政,等.新型冠状病毒PLpro蛋白酶结构与功能的生物信息学分析[J].病毒学报,2021,37(01):43-51.DOI:10.13242/j.cnki.bingduxuebao.003841.
基金信息:
温州市新型冠状病毒肺炎疫情防控应急攻关项目(项目号：ZY202002)，题目：抗新冠状病毒相关诊断治疗产品的研发及产业化; 温州市基础性科研项目(项目号：Y20180059)，题目：宫颈癌靶向治疗制剂ZHPV16E7affitoxin384的临床转化试验前研究;温州市基础性科研项目(项目号：Y20180072)，题目：基于HBcAg纳米颗粒载体的HPV16型E6/E7蛋白多表位治疗性疫苗研究; 温州医科大学科研启动项目(项目号：QTJ14040)，题目：人乳头瘤病毒E7特异性亲和体分子作用机制研究
2020-04-07
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