Production of Recombinant Chicken IgY-Fc and Evaluation of Its Transport Ability into Avian Egg Yolks
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概要
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Maternal immunoglobulin (Ig) Y, the avian counterpart of mammalian IgG, is efficiently transported into the yolks of maturing oocytes. We have previously shown that the Fc region plays a critical role in the IgY transport into avian egg yolks. The aim of this study was to produce recombinant IgY-Fc and to evaluate its transport ability into avian egg yolks. Two basic expression vectors were constructed: one mainly expressed three constant regions of chicken IgY heavy chain (Fcυ2-4) that contained three cysteine residues (C252, C340, C347) involved in inter-heavy chain disulfide bonds; and the other mainly expressed two constant regions (Fcυ3-4) that contained two cysteine residues (C340, C347). SDS-PAGE and Western blotting analyses indicated that both Fcυ2-4 and Fcυ3-4 were separated into multiple protein bands under non-reducing conditions, suggesting incomplete formation of inter-chain disulfide bonds. To prevent the incomplete disulfide bond formation, C340 and C347 were mutated individually to serine by site-directed mutagenesis. As expected, each of the mutated recombinant IgY-Fc produced a single protein band by SDS-PAGE analysis. When intravenously-injected into laying quail, the two mutants, Fcυ2-4C347S and Fcυ3-4C340S, were transported into the egg yolks at high levels, whereas the other two mutants, Fcυ2-4C340S and Fcυ3-4C347S, were transported into the egg yolks at lower levels. In conclusion, we have succeeded in producing a recombinant IgY-Fc retaining a high transport ability into the avian egg yolks; this protein will be provided as a useful tool for studying the mechanism of IgY transport into egg yolks.
- 日本家禽学会の論文
日本家禽学会 | 論文
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