Expression and functional characterization of recombinant tributyltin-binding protein type 2

Hina Satone, Eriko Akahoshi, Aiko Nakamura, Jae Man Lee, Masato Honda, Yohei Shimasaki, Shun ichiro Kawabata, Takahiro Kusakabe, Yuji Oshima

研究成果: ジャーナルへの寄稿レター

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Tributyltin-binding proteins (TBT-bps) are members of the fish lipocalins that were isolated from the blood of Japanese flounder (Paralichthys olivaceus) and function in the binding and detoxification of TBT. In this study, we constructed a baculovirus-silkworm expression system and obtained recombinant TBT-bp2 (rTBT-bp2; 31.5 kDa) from the hemolymph of silkworm larvae injected with a recombinant baculovirus containing the TBT-bp2 gene. The binding potential of rTBT-bp2 was investigated and compared to that of the previously available recombinant TBT-bp1 (rTBT-bp1). Both rTBT-bp2 and rTBT-bpl bound to DAUDA, a typical fluorescent ligand of lipocalins, with dissociation constants of 0.97 and 1.75 μM, respectively. The Hill coefficient value indicated that rTBT-bp2 may have multiple binding sites and strong negative cooperativity. These results suggest that the typical central cavity of lipocalins composed of eight specific P-sheets is conserved in rTBT-bp2, as it is in rTBT-bpl, although rTBT-bp2 has different effects than rTBT-bp1 in TBT binding. In a competition assay, rTBT-bp2 displayed exponential binding affinity to TBT with an inhibition constant of 0.29 μM, demonstrating that TBT binds to the central ligand pocket of rTBT-bp2. However, three fatty acids did not show any affinity to rTBT-bp2. Further studies are required to elucidate the endogenous function of TBT-bps as fish lipoca-lins and their function in responding to xenobiotics.

元の言語英語
ページ(範囲)885-890
ページ数6
ジャーナルJournal of Toxicological Sciences
38
発行部数6
DOI
出版物ステータス出版済み - 11 18 2013

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All Science Journal Classification (ASJC) codes

  • Toxicology

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