木薯果胶甲基酯酶抑制因子MePMEI1的生物信息学分析
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摘要
为探讨木薯MePMEI1的分子结构特征。通过PCR扩增和测序技术及生物信息学分析工具对木薯MePMEI1基因进行克隆、测序及相关生物信息学分析。结果表明木薯MePMEI1基因编码区全长609 bp,编码202个氨基酸残基;MePMEI1基因编码蛋白分子量21.78 k D,理论等电点(pI)约为5.51;生物信息学预测发现,木薯MePMEI1蛋白是稳定的亲水蛋白;具有跨膜区为分泌蛋白;含有1个PMEI结构域,1个糖基化位点,31个磷酸化位点;二、三级结构以α螺旋和无规则卷曲为主。该蛋白的生物功能可能与细胞被膜、酶和生长因子等相关。木薯MePMEI1基因的生物信息学分析为进一步研究其遗传特性和生理生化机制提供了理论依据。
In order to investigate the molecular structure of cassava Me PMEI1, the cassava Me PMEI1 gene was cloned and sequenced by PCR and sequencing technology and analyzed by bioinformatics tools. The results showed that the coding area of cassava Me PMEI1 gene was 609 bp in length, encoding 202 amino acid residues. The molecular weight of Me PMEI1 gene encoding protein was 21.78 k D and the theoretical isoelectric point(pI) was about 5.51. The bioinformatics predicted that the Me PMEI1 protein was stable hydrophilic protein and was secreted protein with transmembrane region. The protein contained one PMEI domain, one glycosylation site, and 31 phosphorylation sites, of which the secondary and tertiary structures were mainly composed of α-helix and random coil. The biological function of the protein might be related to cell membranes, enzymes and growth factors.Bioinformatics analysis of cassava Me PMEI1 gene would lay the foundation for further study of the genetic characteristics and physiological and biochemical mechanisms of this gene.
In order to investigate the molecular structure of cassava Me PMEI1, the cassava Me PMEI1 gene was cloned and sequenced by PCR and sequencing technology and analyzed by bioinformatics tools. The results showed that the coding area of cassava Me PMEI1 gene was 609 bp in length, encoding 202 amino acid residues. The molecular weight of Me PMEI1 gene encoding protein was 21.78 k D and the theoretical isoelectric point(pI) was about 5.51. The bioinformatics predicted that the Me PMEI1 protein was stable hydrophilic protein and was secreted protein with transmembrane region. The protein contained one PMEI domain, one glycosylation site, and 31 phosphorylation sites, of which the secondary and tertiary structures were mainly composed of α-helix and random coil. The biological function of the protein might be related to cell membranes, enzymes and growth factors.Bioinformatics analysis of cassava Me PMEI1 gene would lay the foundation for further study of the genetic characteristics and physiological and biochemical mechanisms of this gene.
引文
An S.H.,S ohn K.H.,Choi H.W.,Hwang I.S.,Lee S.C.,and Hwang B.K.,2008,Pepper pectin methylesterase inhibitor protein CaPMEI1 is required for antifungal activity,basal disease resistance and abiotic stress tolerance,Planta,228(1):61-78
Bellincampi D.,Cervone F.,and Lionetti V.,2014,Plant cell wall dynamics and wall-related susceptibility in plant-pathogen interactions,Front.Plant Sci.,5:228
Blom N.,Gammeltoft S.,and Brunak S.,1999,Sequence and structure-based prediction of eukaryotic protein phosphorylation sites,Journal of Molecular Biology,294(5):1351-1362
Di Matteo A.,Giovane A.,Raiola A.,Camardella L.,Bonivento D.,De Lorenzo G.,Cervone F.,Bellincampi D.,and Tsernoglou D.,2005,Structural basis for the interaction between pectin methylesterase and a specific inhibitor protein,Plant Cell,17(3):849-858
Gasteiger E.,Hoogland C.,Gattiker A.,Duvaud S.,Wilkins M.R.,Appel R.D.,and Bairoch A.,2005,Protein identification and analysis tools on the ExPASy server,In:Walker J.M.(ed.),The proteomics protocols handbook,Humana Press,Totowa,New Jersey,pp.571-607
Harholt J.,Suttangkakul A.,and Scheller H.V.,2010,Biosynthesis of pectin,Plant Physiol,153(2):384-395
Herron S.R.,Benen J.A.,Scavetta R.D.,Visser J.,and Jurnak F.,2000,Structure and function of pectic enzymes:virulence factors of plant pathogens,Proc.Natl.Acad.Sci.USA,97(16):8762-8769
Hongo S.,Sato K.,Yokoyama R.,and Nishitani K.,2012,Demethylesterification of the primary wall by PECTINMETHYLESTERASE35 provides mechanical support to the Arabidopsis stem,Plant Cell,24(6):2624-2634
Jensen L.J.,Gupta R.,Blom N.,Devos D.,Tamames J.,Kesmir C.,Nielsen H.,Strfeldt H.H.,Rapacki K.,Workman C.,Andersen C.A.F.,Knudsen S.,Krogh A.,Valencia A.,and Brunak S.,2002,Ab initio prediction of human orphan protein function from post-translational modifications and localization features,J.Mol.Biol.,319:1257-1265
Jeong H.Y.,Nguyen H.P.,and Lee C.,2014,Genome-wide identification and expression analysis of rice pectin methylesterases:implication of functional roles of pectin modification in rice physiology,J.Plant Physiol.,183:23-29
Jithesh M.N.,Wally O.D.S.,Manfield I.,Critchley A.T.,Hiltz D.,and Prithiviraj B.,2012,Analysis of seaweed extract-induced transcriptome leads to identification of a negative regulator of salt tolerance in Arabidopsis,Hort Science,47(6):704-709
Kelley L.A.,Mezulis S.,Yates C.M.,Wass M.N.,and Sternberg M.J.,2015,The Phyre2 web portal for protein modeling,prediction and analysis,Nature Protocols,10(6):845-858
Kim S.J.,Held M.A.,Zemelis S.,Wilkerson C.,and Brandizzi F.,2015,CGR2 and CGR3 have critical overlapping roles in pectin methylesterification and plant growth in Arabidopsis thaliana,Plant J.,82(2):208-220
Lionetti V.,Cervone F.,and Bellincampi D.,2012,Methyl esterification of pectin plays a role during plant-pathogen interactions and affects plant resistance to diseases,J.Plant Physiol.,169(16):1623-1630
Lionetti V.,Fabri E.,De Caroli M.,Hansen A.R.,Willats W.G.T.,Piro G.,and Bellincampi D.,2017,Three pectin methyl esterase inhibitors protect cell wall integrity for immunity to Botrytis,Plant Physiol.,173:1844-1863
Lionetti V.,Raiola A.,Cervone F.,and Bellincampi D.,2013,Transgenic expression of pectin methylesterase inhibitors limits tobamovirus spread in tobacco and Arabidopsis,Mol.Plant Pathol.,15(3):265-274
Lionetti V.,Raiola A.,Mattei B.,and Bellincampi D.,2015,The grapevine Vv PMEI1 gene encodes a novel functional pectin methylesterase inhibitor associated to grape berry development,PLoS One,10(7):e0133810
Müller K.,Levesque-Tremblay G.,Bartels S.,Weitbrecht K.,Wormit A.,Usadel B.,Haughn G.,and Kermode A.R.,2013,Demethylesterification of cell wall pectins in Arabidopsis plays a role in seed germination,Plant Physiol.,161(1):305-316
Nguyen H.P.,Jeong H.Y.,Jeon S.H.,Kim D.,and Lee C.,2017,Rice pectin methylesterase inhibitor28(Os PMEI28)encodes a functional PMEI and its overexpression results in a dwarf phenotype through increased pectin methylesterification levels,J.Plant Physiol.,208:17-25
Nielsen H.,2017,Predicting secretory proteins with SignalP,In:Kihara D.(ed.),Methods in molecular biology,Springer,Springe,Germany,pp.59-73
Peaucelle A.,Louvet R.,Johansen J.N.,Hofte H.,Laufs P.,Pelloux J.,and Mouille G.,2008,Arabidopsis phyllotaxis is controlled by the methyl-esterification status of cell-wall pectins,Curr.Biol.,18(24):1943-1948
Pelletier S.,Van Orden J.,Wolf S.,Vissenberg K.,Delacourt J.,Ndong Y.A.,Pelloux J.,Bischoff V.,Urbain A.,Mouille G.,Lemonnier G.,Renou J.P.,and H觟fte H.,2010,A role for pectin de-methylesterification in a developmentally regulated growth acceleration in dark-grown Arabidopsis hypocotyls,New Phytol.,188(3):726-739
Pinzón-Latorre D.,and Deyholos M.K.,2013,Characterization and transcript profiling of the pectin methylesterase(PME)and pectin methylesterase inhibitor(PMEI)gene families in flax(Linum usitatissimum),BMC Genomics,14:742
Raiola A.,Lionetti V.,Elmaghraby I.,Immerzeel P.,Mellerowicz E.J.,Salvi G.,Cervone F.,and Bellincampi D.,2011,Pectin methylesterase is induced in Arabidopsis upon infection and is necessary for a successful colonization by necrotrophic pathogens,Mol.Plant Microbe Interact,24(4):432-440
Saez-Aguayo S.,Ralet M.C.,Berger A.,Botran L.,Ropartz D.,Marion-Poll A.,and North H.M.,2013,PECTIN METHYL-ESTERASE INHIBITOR6 promotes Arabidopsis mucilage release by limiting methylesterification of homogalacturonan in seed coat epidermal cells,Plant Cell,25(1):308-323
Sapay N.,Guermeur Y.,and Deléage G.,2006,Prediction of amphipathic in-plane membrane anchors in monotopic proteins using a SVM classifier,BMC Bioinformatics,7(1):255
Sénéchal F.,Wattier C.,Rustérucci C.,and Pelloux J.,2014,Homogalacturonan-modifying enzymes:structure,expression,and roles in plants,J.Exp.Bot.,65(18):5125-5160
Volpi C.,Raiola A.,Janni M.,Gordon A.,O'Sullivan D.M.,Favaron F.,and D'Ovidio R.,2013,Claviceps purpurea expressing polygalacturonases escaping PGIP inhibition fully infects Pv PGIP2 wheat transgenic plants but its infection is delayed in wheat transgenic plants with increased level of pectin methyl esterification,Plant Physiol.,Biochem.,73:294-301
Wang M.,Yuan D.,Gao W.,Li Y.,Tan J.,and Zhang X.,2013,A comparative genome analysis of PME and PMEI families reveals the evolution of pectin metabolism in plant cell walls,PLoS One,8(8):e72082
Wolf S.,Mravec J.,Greiner S.,Mouille G.,and H觟fte H.,2012,Plant cell wall homeostasis is mediated by brassinosteroid feedback signaling,Curr.Biol.,22(18):1732-1737
Woriedh M.,Wolf S.,Márton M.L.,Hinze A.,Gahrtz M.,Becker D.,and Dresselhaus T.,2013,External application of gametophyte-specific Zm PMEI1 induces pollen tube burst in maize,Plant Reprod.,26(3):255-266
Wydra K.,and Berl H.,2006,Structural changes of homogalacturonan,rhamnogalacturonan I and arabinogalactan protein in xylem cell walls of tomato genotypes in reaction to Ralstonia solanacearum,Physiol.Mol.Plant Pathol.,68(1-3):41-50
Xie Q.F.,Zhao X.D.,Zhang Q.,Huang L.,Mao H.W.,and Zhao Y.,2009,Prediction of N-Glycosylation sites of human metapneumovirus fusion protein and construction of various mutants with modified N-Glycosylation,Shiyong Erke Linchuang Zazhi(Journal of Applied Clinical Pediatrics),24(22):1714-1716)(谢琴芳,赵晓东,张琴,黄路,毛华伟,赵耀,2009,人偏肺病毒融合蛋白N-糖基化位点的预测及不同N-糖基化修饰突变体的构建,实用儿科临床杂志,24(22):1714-1716)
Bellincampi D.,Cervone F.,and Lionetti V.,2014,Plant cell wall dynamics and wall-related susceptibility in plant-pathogen interactions,Front.Plant Sci.,5:228
Blom N.,Gammeltoft S.,and Brunak S.,1999,Sequence and structure-based prediction of eukaryotic protein phosphorylation sites,Journal of Molecular Biology,294(5):1351-1362
Di Matteo A.,Giovane A.,Raiola A.,Camardella L.,Bonivento D.,De Lorenzo G.,Cervone F.,Bellincampi D.,and Tsernoglou D.,2005,Structural basis for the interaction between pectin methylesterase and a specific inhibitor protein,Plant Cell,17(3):849-858
Gasteiger E.,Hoogland C.,Gattiker A.,Duvaud S.,Wilkins M.R.,Appel R.D.,and Bairoch A.,2005,Protein identification and analysis tools on the ExPASy server,In:Walker J.M.(ed.),The proteomics protocols handbook,Humana Press,Totowa,New Jersey,pp.571-607
Harholt J.,Suttangkakul A.,and Scheller H.V.,2010,Biosynthesis of pectin,Plant Physiol,153(2):384-395
Herron S.R.,Benen J.A.,Scavetta R.D.,Visser J.,and Jurnak F.,2000,Structure and function of pectic enzymes:virulence factors of plant pathogens,Proc.Natl.Acad.Sci.USA,97(16):8762-8769
Hongo S.,Sato K.,Yokoyama R.,and Nishitani K.,2012,Demethylesterification of the primary wall by PECTINMETHYLESTERASE35 provides mechanical support to the Arabidopsis stem,Plant Cell,24(6):2624-2634
Jensen L.J.,Gupta R.,Blom N.,Devos D.,Tamames J.,Kesmir C.,Nielsen H.,Strfeldt H.H.,Rapacki K.,Workman C.,Andersen C.A.F.,Knudsen S.,Krogh A.,Valencia A.,and Brunak S.,2002,Ab initio prediction of human orphan protein function from post-translational modifications and localization features,J.Mol.Biol.,319:1257-1265
Jeong H.Y.,Nguyen H.P.,and Lee C.,2014,Genome-wide identification and expression analysis of rice pectin methylesterases:implication of functional roles of pectin modification in rice physiology,J.Plant Physiol.,183:23-29
Jithesh M.N.,Wally O.D.S.,Manfield I.,Critchley A.T.,Hiltz D.,and Prithiviraj B.,2012,Analysis of seaweed extract-induced transcriptome leads to identification of a negative regulator of salt tolerance in Arabidopsis,Hort Science,47(6):704-709
Kelley L.A.,Mezulis S.,Yates C.M.,Wass M.N.,and Sternberg M.J.,2015,The Phyre2 web portal for protein modeling,prediction and analysis,Nature Protocols,10(6):845-858
Kim S.J.,Held M.A.,Zemelis S.,Wilkerson C.,and Brandizzi F.,2015,CGR2 and CGR3 have critical overlapping roles in pectin methylesterification and plant growth in Arabidopsis thaliana,Plant J.,82(2):208-220
Lionetti V.,Cervone F.,and Bellincampi D.,2012,Methyl esterification of pectin plays a role during plant-pathogen interactions and affects plant resistance to diseases,J.Plant Physiol.,169(16):1623-1630
Lionetti V.,Fabri E.,De Caroli M.,Hansen A.R.,Willats W.G.T.,Piro G.,and Bellincampi D.,2017,Three pectin methyl esterase inhibitors protect cell wall integrity for immunity to Botrytis,Plant Physiol.,173:1844-1863
Lionetti V.,Raiola A.,Cervone F.,and Bellincampi D.,2013,Transgenic expression of pectin methylesterase inhibitors limits tobamovirus spread in tobacco and Arabidopsis,Mol.Plant Pathol.,15(3):265-274
Lionetti V.,Raiola A.,Mattei B.,and Bellincampi D.,2015,The grapevine Vv PMEI1 gene encodes a novel functional pectin methylesterase inhibitor associated to grape berry development,PLoS One,10(7):e0133810
Müller K.,Levesque-Tremblay G.,Bartels S.,Weitbrecht K.,Wormit A.,Usadel B.,Haughn G.,and Kermode A.R.,2013,Demethylesterification of cell wall pectins in Arabidopsis plays a role in seed germination,Plant Physiol.,161(1):305-316
Nguyen H.P.,Jeong H.Y.,Jeon S.H.,Kim D.,and Lee C.,2017,Rice pectin methylesterase inhibitor28(Os PMEI28)encodes a functional PMEI and its overexpression results in a dwarf phenotype through increased pectin methylesterification levels,J.Plant Physiol.,208:17-25
Nielsen H.,2017,Predicting secretory proteins with SignalP,In:Kihara D.(ed.),Methods in molecular biology,Springer,Springe,Germany,pp.59-73
Peaucelle A.,Louvet R.,Johansen J.N.,Hofte H.,Laufs P.,Pelloux J.,and Mouille G.,2008,Arabidopsis phyllotaxis is controlled by the methyl-esterification status of cell-wall pectins,Curr.Biol.,18(24):1943-1948
Pelletier S.,Van Orden J.,Wolf S.,Vissenberg K.,Delacourt J.,Ndong Y.A.,Pelloux J.,Bischoff V.,Urbain A.,Mouille G.,Lemonnier G.,Renou J.P.,and H觟fte H.,2010,A role for pectin de-methylesterification in a developmentally regulated growth acceleration in dark-grown Arabidopsis hypocotyls,New Phytol.,188(3):726-739
Pinzón-Latorre D.,and Deyholos M.K.,2013,Characterization and transcript profiling of the pectin methylesterase(PME)and pectin methylesterase inhibitor(PMEI)gene families in flax(Linum usitatissimum),BMC Genomics,14:742
Raiola A.,Lionetti V.,Elmaghraby I.,Immerzeel P.,Mellerowicz E.J.,Salvi G.,Cervone F.,and Bellincampi D.,2011,Pectin methylesterase is induced in Arabidopsis upon infection and is necessary for a successful colonization by necrotrophic pathogens,Mol.Plant Microbe Interact,24(4):432-440
Saez-Aguayo S.,Ralet M.C.,Berger A.,Botran L.,Ropartz D.,Marion-Poll A.,and North H.M.,2013,PECTIN METHYL-ESTERASE INHIBITOR6 promotes Arabidopsis mucilage release by limiting methylesterification of homogalacturonan in seed coat epidermal cells,Plant Cell,25(1):308-323
Sapay N.,Guermeur Y.,and Deléage G.,2006,Prediction of amphipathic in-plane membrane anchors in monotopic proteins using a SVM classifier,BMC Bioinformatics,7(1):255
Sénéchal F.,Wattier C.,Rustérucci C.,and Pelloux J.,2014,Homogalacturonan-modifying enzymes:structure,expression,and roles in plants,J.Exp.Bot.,65(18):5125-5160
Volpi C.,Raiola A.,Janni M.,Gordon A.,O'Sullivan D.M.,Favaron F.,and D'Ovidio R.,2013,Claviceps purpurea expressing polygalacturonases escaping PGIP inhibition fully infects Pv PGIP2 wheat transgenic plants but its infection is delayed in wheat transgenic plants with increased level of pectin methyl esterification,Plant Physiol.,Biochem.,73:294-301
Wang M.,Yuan D.,Gao W.,Li Y.,Tan J.,and Zhang X.,2013,A comparative genome analysis of PME and PMEI families reveals the evolution of pectin metabolism in plant cell walls,PLoS One,8(8):e72082
Wolf S.,Mravec J.,Greiner S.,Mouille G.,and H觟fte H.,2012,Plant cell wall homeostasis is mediated by brassinosteroid feedback signaling,Curr.Biol.,22(18):1732-1737
Woriedh M.,Wolf S.,Márton M.L.,Hinze A.,Gahrtz M.,Becker D.,and Dresselhaus T.,2013,External application of gametophyte-specific Zm PMEI1 induces pollen tube burst in maize,Plant Reprod.,26(3):255-266
Wydra K.,and Berl H.,2006,Structural changes of homogalacturonan,rhamnogalacturonan I and arabinogalactan protein in xylem cell walls of tomato genotypes in reaction to Ralstonia solanacearum,Physiol.Mol.Plant Pathol.,68(1-3):41-50
Xie Q.F.,Zhao X.D.,Zhang Q.,Huang L.,Mao H.W.,and Zhao Y.,2009,Prediction of N-Glycosylation sites of human metapneumovirus fusion protein and construction of various mutants with modified N-Glycosylation,Shiyong Erke Linchuang Zazhi(Journal of Applied Clinical Pediatrics),24(22):1714-1716)(谢琴芳,赵晓东,张琴,黄路,毛华伟,赵耀,2009,人偏肺病毒融合蛋白N-糖基化位点的预测及不同N-糖基化修饰突变体的构建,实用儿科临床杂志,24(22):1714-1716)