文章摘要
王艳华,张亦陈,孙妍,刘逸尘,耿绪云,孙金生.中华绒螯蟹高血糖激素基因的克隆和分子结构解析[J].水产学报,2013,37(7):987~993
中华绒螯蟹高血糖激素基因的克隆和分子结构解析
Cloning and molecular structure analysis of crustacean hyperglycemic hormone(Ers-CHH)in Eriocheir sinensis
投稿时间:2012-11-15  修订日期:2013-03-28
DOI:10.3724/SP.J.1231.2013.38432
中文关键词: 中华绒螯蟹  高血糖激素  克隆  分子结构
英文关键词: Eriocheir sinensis  crustacean hyperglycemic hormone  clone  molecular structure
基金项目:国家“八六三”高技术研究发展计划(2012AA10A401);国家科技支撑计划项目(2011BAD13B07);国家自然科学基金项目(30871907);天津市应用基础研究重点项目(10JCZDJC18200)
作者单位
王艳华 天津师范大学生命科学学院,天津市动植物抗性重点实验室 
张亦陈 天津师范大学生命科学学院,天津市动植物抗性重点实验室 
孙妍 天津市水生动物疫病预防控制中心 
刘逸尘 天津师范大学生命科学学院,天津市动植物抗性重点实验室 
耿绪云 天津市水生动物疫病预防控制中心 
孙金生 天津师范大学生命科学学院,天津市动植物抗性重点实验室 
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中文摘要:
      为深入研究中华绒螯蟹高血糖激素(crustacean hyperglycemic hormone,CHH)基因的结构与功能,首次从中华绒螯蟹眼柄中克隆得到高血糖激素基因的全长cDNA序列(Ers-CHH,GenBank 登录号:JX485664)。序列及分子结构分析结果显示:Ers-CHH基因全长585 bp,ORF区为 453 bp,编码150 个氨基酸,N端30个氨基酸为预测的信号肽区域,其后为42个氨基酸的前体相关肽和78 个氨基酸的成熟肽。其成熟肽区具有典型的CHH家族结构域,包括3个二硫键C7-C43、C23-C39、C26-C52和4个α螺旋(alpha-helix),但不具有β折叠结构(beta-sheet)。Ers-CHH成熟肽氨基酸序列与溪蟹相比有92%的相似性;其三维结构与其它物种CHH三维结构比较结果显示,4个α螺旋所处的位置及所含氨基酸的相似性较高。与中华绒螯蟹MIH相比,CHH序列前段缺少一个螺旋,但其4个α螺旋的位置及氨基酸数目与MIH的α2-α5螺旋高度一致,空间结构也基本相同,这可能是两种激素功能部分重叠的分子基础。上述结果为进一步研究CHH在中华绒螯蟹生长发育及能量代谢过程中的调节机制和人工控制甲壳动物生长代谢提供了重要信息。
英文摘要:
      Crustacean hyperglycemic hormone(CHH)is a peptide hormone originally identified in a crustacean neurosecretory complex,the X-organ/sinus gland complex,and located within the eyestalks.CHH is placed in the crustacean hyperglycemic hormone family,which also includes molt-inhibiting hormone(MIH),vitellogenesis-inhibiting hormone(VIH)or gonad-inhibiting hormone(GIH),mandibular organ-inhibiting hormone(MOIH)and insect ion transport peptide(ITP).CHH plays a significant role in the growth,molting and reproduction of crustaceans.By using degenerate primers which were designed according to the N-terminal amino acid sequences of CHH isolated by our lab and the sequences of homologous species CHH,a novel cDNA of crustacean hyperglycemic hormone gene(Ers-CHH,GenBank Accession No:JX485664)was cloned from Chinese mitten crab(Eriocheir sinensis).The full length cDNA consists of 585 bp with a 453 bp open reading frame,encoding 150 amino acids,containing a 30 amino acids signal peptide,and a 42 amino acids CHH precursor-related peptide,a 78 amino acids mature peptide.The mature peptide contains 6 conserved cysteines which formed three disulfide bonds C7-C43、C23-C39、C26-C52.There was an arthropod CHH/MIH/GIH neurohormones family signature in this domain.The three-dimensional structure prediction showed that Ers-CHH mature peptide consisted of 4 α-helixes,and not with the β-sheet structure.5 Prosite patterns,which were analyzed by a Swiss-PdbViewer(v4.0.4),are PS00005:Protein kinase C phosphorylation site(Ser6,Lys8);PS00006:Casein kinase Ⅱ phosphorylation site(Ser17,Glu20);PS00008:N myristoylation site(Gly38,Cys39,Asn42,Cys43);PS00342:Microbodies C-terminal targeting signal(Ser17,Lys18,Leu19);PS01250:Arthropod CHH/MIH/GIH neurohormones family(Val35,Cys39,Arg40,Asn42,Cys43,Tyr44,Asn46,Phe49,Cys52).The results of the structural model comparison show that the structure of Ers-CHH has high similarity with other CHHs and Ers-CHH.They have no β-sheet structure,except that CHH has 4α-helixes,MIH has 5α-helixes.Although α1 helix of MIH could not be found in CHHs,the position and the amino acid number of α2-α5 helixes in MIH molecular were same as the α1-α4 helixes in CHH.It might be able to interpret the function similarity between CHH and MIH.Multiple alignment results showed that Ers-CHH has the highest identity with Ptychognathus pusillus CHH(92%),and it also shared high identities with Neohelice granulata(86%)and Pachygrapsus marmoratus(72%).However,it showed lower identity with CHH from crayfishes,such as Nephrops norvegicus(49%)and Homarus gammarus(46%).Multiple alignments of Ers-CHH and MIH from other crustaceans showed that only six amino acid residues(Arg13,Asp25,Asn28,Arg31,Arg40,Phe49)in Ers-CHH sequence were same with MIH sequences.It may be helpful for further research on regulation mechanism of CHH in the process of crab growth and carbohydrate metabolism.
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