Purification and characterization of tributyltin-binding protein of tiger puffer, Takifugu rubripes

We successfully purified Trub.TBT-bp alpha, a tributyltin (TBT) binding protein (bp) of the tiger puffer. Takifugu rubripes. Tiger puffer was injected intraperitoneally with TBT (1.0 mg/kg body weight) and Trub.TBT-bp alpha was purified from serum by ammonium sulfate fractionation, gel filtration chromatography and polyacrylamide gel electrophoresis. Gel electrophoresis revealed that the Trub.TBT-bp alpha has a molecular mass of approximately 48.5 kDa and contains at least 40% N-glycan. The deduced 212 amino acid sequence of the protein showed the highest identity (41%, 212 amino acid overlap and E-value: 9e-42) with TBT-binding protein type 1 (TBT-bp1) of Paralichthys olivaceus (Japanese flounder). Analysis of the gene structure of Trub.TBT-bp alpha suggests that this protein belongs to the lipocalin superfamily, which may be important in the accumulation and elimination of TBT. Phylogenetic analysis suggests that functionalization of TBT-bps has occurred during evolution, and that the functions of this group of proteins might be important for fish survival. (C) 2010 Elsevier Inc. All rights reserved.