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重组人神经肽Y受体Y5融合蛋白的表达、纯化及其生物信息学分析研究 被引量：1

Study on expression,purification and bioinformatic analysis of human neuropeptideY Y5 receptor
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摘要 目的在大肠杆菌中表达人神经肽YY5受体,并对之进行纯化、鉴定及生物信息学分析。方法取已构建好且经测序确认无误的重组质粒pET28a-Y5转化大肠杆菌BL21(DE3),IPTG诱导表达融合蛋白,并经SDS-PAGE检测和Western印迹鉴定,表达产物包涵体经Ni2+-NTA亲和层析纯化。然后利用相关在线软件进行生物信息学分析Y5受体蛋白。结果经IPTG诱导含有pET28a-Y5重组质粒的DE3菌,表达出重组人Y5融合蛋白。重组蛋白经Ni2+-NTA亲和层析进行纯化后,得到了较高纯度的融合蛋白。经相关在线软件分析后获得了Y5受体的相关生物学特性。结论重组质粒pET28a-Y5在大肠杆菌DE3中成功表达,亲和层析纯化后获得较高纯度融合蛋白,并对Y5受体蛋白的生物学特征进行了预测,为进一步研究其生物学功能及其抗体的研制奠定了基础。 Objective To express human Y5 receptor protein in E.coli,purify and identify it,and conduct bioinformatic analysis of Y5 receptor.Methods The recombinant plasmid pET28a-Y5 which had been well constructed and sequentially confirmed was transplanted into E.coli BL21（DE3） and induced by IPTG to express fusion proteins.SDS-PAGE and Western blot were used to test and identify the expressed fusion proteins.The inclusion body of the expressed product was purified by Ni2＋-NTA affinity chromatography.Then bioinformatic analysis of the Y5 receptor protein was conducted with the help of related online software.Results After being induced by IPTG,the DE3 with recombinant plasmid pET28a-Y5 expressed recombinant human Y5 receptor protein.Highly purified Y5 receptor fusion protein was obtained by Ni^2＋-NTA affinity chromatography.Related biological characteristics of Y5 receptor were obtained after the online software analysis.Conclusions The recombinant plasmid pET28a-Y5 can be successfully expressed in DE3.Highly purified proteins can be obtained by Ni2＋-NTA affinity chromatography.Y5 receptor＇s biological characteristics are predicted,which lays foundation for further studies of Y5 receptor＇s biological function and antibody development.
作者 丁克祥 董萍 郑永晨 杨永鹏 朱晓亮 韩晋云 单志新 丁宇 丁振华
机构地区 南方医科大学科研部 大连董萍美容整形医院 吉林大学附属第二临床医院 南方医科大学附属南方医院 南方医科大学研究生学院 南方医科大学附属华南医院 华中科技大学同济医学院附属同济医院 南方医科大学公共卫生与热带医学学院
出处 《中国老年学杂志》 CAS CSCD 北大核心 2009年第20期2609-2613,共5页 Chinese Journal of Gerontology
关键词 人神经肽YY5受体 融合蛋白 包涵体 纯化 生物信息学 NeuropeptideY Y5 receptor Fusion protein Inclusion body Purification Bioinformatics
分类号 R392.4 [医药卫生—免疫学] R341.6 [医药卫生—基础医学]
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参考文献10

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二级参考文献8

  • 1MacNeil, Douglas J. NPY Y1 and Y5 Receptor selective antagonists as anti-obesity drugs [J] Medicinal Chemistry, 2007, 7:1721-1733.
  • 2Gerald C, Walker MW, Criscionc L, ct al. A receptor subtype involved in neuropeptide Y - induced food intake [J] . Nature, 1996; 382: 168 -171.
  • 3Tang - Christensen M, Kristensen P, Stidsen CE, et al. Central administration of Y5receptor antisensedcoreasess spontaneous food intake and attenuatesfeeding in response to exogenous neuropeptide Y [J] . J Endocrinol, 1998; 159:307-312.
  • 4Schaflhauser AO, Stricker - Krongrad A, Brunner L, et al. Inhibition of foodintake by neuropeptideY Y5 receptor antisense oligodeoxynucleotides [J] . Ddiabetes, 1997, 46 (11): 1792-1798.
  • 5Sato N, Takahashi T, Shibata T, et al. Design and synthesis of the potent, orally available, braipenetrable arylpyrasole class of neuropeptide Y5 receptor antagonists [J] . J Med Chem, 2003, 46:666 -669.
  • 6Wyss P, Stricker - Krongrad A, Brunner L, et al. The pharlnacology of neuropeptideY (NpY) receptor mediated feedingin rats characterizes betterY5 thanY1, but notY2 or Y4 subtypes [J] . Regul PePt, 1998, 75: 363-371.
  • 7Mashiko S, Ishihara A, Iwaasa H, et aL Characterization of neuropeptideY (NPY) Y5 receptor- mediated obesity in mice: chronic intracerebroventricula rlnfusionof of D - Trp34NPY [J] . Endocrinology, 2003, 144: 1793 - 1801.
  • 8Ishihara A, Kanatani A, Mashiko S, et al. A neuropeptide Y Y5 angagonist selectively ameliorates body weight gain and assoeiatde parameters in diet - induced obese mice[ J]. PNAS,2006,103:7154 -7158.

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中国老年学杂志
2009年 第20期

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