| HAN Yinglun,GOU Meng,SONG Xiaoping,SONG Tao,SHI Biyue,PANG Yue,LI Qingwei. 2018. Target-directed isolation and identification of a serum lectin from lamprey (Lampetra japonica) by chromatographys and MALDI-TOF/TOF. Acta Oceanologica Sinica, 37(5):113－116 |
| Target-directed isolation and identification of a serum lectin from lamprey (Lampetra japonica) by chromatographys and MALDI-TOF/TOF |
| 利用色谱和飞行时间质谱方法分离并鉴定七鳃鳗血清中凝集素 |
| Received:March 13, 2016 Revised:December 29, 2017 |
| DOI：10.1007/s13131-018-1175-7 |
| Key words:Lampetra japonica lectin purification identification MALDI-TOF/TOF |
| 中文关键词: 七鳃鳗 凝集素 纯化 鉴定 飞行时间质谱 |
| 基金项目:The National Program on Key Basic Research Project (973 Program) of China under contract No. 2013CB835304; the National Marine Public Projects under contract No. 201305016; the National Natural Science Foundation of China under contract Nos 31772884 and 31601865; the Key Projects of Scientific Research Platform of Liaoning Provincial Education Department under contract No. L201683651. |
| Author Name | Affiliation | E-mail | | HAN Yinglun | College of Life Science, Liaoning Normal University, Dalian 116029, China
Lamprey Research Center, Liaoning Normal University, Dalian 116029, China | | | GOU Meng | College of Life Science, Liaoning Normal University, Dalian 116029, China
Lamprey Research Center, Liaoning Normal University, Dalian 116029, China | | | SONG Xiaoping | College of Life Science, Liaoning Normal University, Dalian 116029, China
Lamprey Research Center, Liaoning Normal University, Dalian 116029, China
Affiliated Zhongshan Hospital of Dalian University, Dalian 116001, China | | | SONG Tao | College of Life Science, Liaoning Normal University, Dalian 116029, China
Lamprey Research Center, Liaoning Normal University, Dalian 116029, China | | | SHI Biyue | College of Life Science, Liaoning Normal University, Dalian 116029, China
Lamprey Research Center, Liaoning Normal University, Dalian 116029, China | | | PANG Yue | College of Life Science, Liaoning Normal University, Dalian 116029, China
Lamprey Research Center, Liaoning Normal University, Dalian 116029, China | | | LI Qingwei | College of Life Science, Liaoning Normal University, Dalian 116029, China
Lamprey Research Center, Liaoning Normal University, Dalian 116029, China | liqw@263.net |
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| Abstract: |
| A 105-kDa polymer lectin was purified from lamprey (Lampetra japonica) serum by chromatography methods including cation ion-exchange chromatography with a SP-SepharoseTM XL column and size exclusion chromatography with a Superdex 200 column. The target fractions were collected according to the direction of hemagglutinating activity. The results revealed that the active fractions could adsorb on SP-Sepharose column and showed a 280 nm UV absorbance peak corresponding to molecular weights of 105 kDa in the following size exclusion chromatography. The target fractions with hemagglutinating activity were further checked by Native-PAGE and SDS-PAGE. Two single bands at around 105 kDa and 35 kDa were displayed by two electrophoresis methods respectively, indicating that the protein exists as a trimer in solution. After Native-PAGE and SDS-PAGE, two bands were excised from the gels respectively and further identified by MALDI-TOF/TOF as serum lectin (gi:13094239). The lectin was able to agglutinate rabbit red blood cells (RRBCs) and sheep red blood cells (SRBCs) in vitro. The lectin isolated from lamprey serum in the current study might be helpful for deeply understanding the innate immune molecules dependent immune defence in jawless vertebrates which have been proved recently that they possess a lymphocyte-based system of anticipatory immunity with variable lymphocyte receptors as mediators. |
| 中文摘要: |
| 本研究利用阳离子交换和分子筛的方法成功分离鉴定出分子量为105kDa，以多聚体形式存在七鳃鳗凝集素。实验结果表明，首先运用阳离子交换色谱方法能够将七鳃鳗血清混合组分进行初步分离，结合凝集活性鉴定方法，鉴定凝集活性最强组分存在于第二峰中。其次；利用分子筛方法将具有凝集活性组分进一步分离，最终得到分子量为105kDa左右具有凝集活性组分。结合Native-PAGE和SDS-PAGE结果表明，凝集活性组分以三聚体形式存在。同时，飞行时间质谱方法鉴定结果为凝集素（gi:13094239）。最终，体外鉴定结果表明七鳃鳗凝集素对兔红细胞和山羊红细胞均具有较强的凝集活性。七鳃鳗凝集素的分离纯化对研究七鳃鳗先天免疫防御机制和可变淋巴受体介导的适应性免疫防御机制均具有重要意义。 |
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