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https://tkuir.lib.tku.edu.tw/dspace/handle/987654321/120052
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| 題名: | Simultaneous Separation of Taxon-Specific Crystallins from Mule Duck and Characterization of Their Enzymatic Activities and Structures |
| 作者: | Wang, Chih-Hsien;Huang, Chia-Chi;Chen, Wenlung |
| 關鍵詞: | Taxon-specific protein;Ion-exchange chromatography;ɛ-Crystallin;δ-Crystallin;FT-Raman;Enzyme activity |
| 日期: | 2017-05-17 |
| 上傳時間: | 2021-03-05 12:14:22 (UTC+8) |
| 摘要: | Methods to obtain pure proteins in large amounts are indispensible in protein research. We report here a large-scale/simultaneous isolation of taxon-specific crystallins (ɛ- and δ-crystallin) from the eye lenses of Mule duck. We also investigate the compositions, enzymatic activities, and structures of these purified taxon-specific proteins. A relatively mild method of ion-exchange chromatography was developed to fractionate ɛ-crystallin and δ-crystallin in large amount, ca. ∼6.60 mg/g-lens and ∼41.0 mg/g-lens, respectively. Both crystallins were identified by electrophoresis, HPLC, and MALDI-TOF-MS. ɛ-Crystallin, with native composition of Mr 142 kDa, consisted of two subunits of 35 kDa and 36 kDa, while δ-Crystallin, with native molecular mass of 200 kDa, comprised single subunit of Mr ∼50 kDa. Both ɛ- and δ-crystallin were tetramers. The former showed lactate dehydrogenase (LDH) activity, while the latter appeared slightly active in an argininosuccinate lyase (ASL) assay. Raman spectroscopic results indicated that the secondary structures of ɛ- and δ-crystallin were predominantly α-helix as evidenced by the vibrational stretching of amide III over 1260 cm−1 and amide I at 1255 cm−1, in greatly contrast to the anti-parallel β-sheet of α- and β-crystallin as demonstrated by amide III at 1238 cm−1 and amide I at 1672 cm−1. The microenvironments of aromatic amino acids and the status of thiol groups also vary in different crystallins. The compositions, enzyme activities, and structures of the ɛ- and δ-crystalline of Mule duck are different from those of Muscovy duck (Cairina moschata) or Kaiya duck (Anas Platyrhynchos var. domestica), which reflect faithfully species specificity. |
| 關聯: | Journal of Chromatography B 1053, p.34-41 |
| DOI: | 10.1016/j.jchromb.2017.02.027 |
| 顯示於類別: | [化學學系暨研究所] 期刊論文
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