Ligandinの構造と機能に関する研究

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Rat liver ligandin was purified to homogeneity by TEAE-cellulose, Sephadex G-75 and QAE Sephadex A-50 chromatography and consisted of equal amounts of two subunits of 22,000(Ya) and 25,000(Yb) daltons. Ya and Yb were also purified to homogeneity by the method of preparative SDS-polyacrylamide gel electrophoresis in a Tris-glycine discontinuous buffer system. Amino acid analysis of Ya, Yb and liver ligandin revealed that the two subunits awere closely related except that most of the cysteine residues of ligandin were on Yb. Ya and Yb had immunological reactivity with liver ligandin. The relative proportion of subunit Ya increased when liver ligandin was stored at -20℃ for a long time. The Ya homodimer was purified by Thiol-Sepharose affinity and QAE Sephadex A-50 chromatography. More than 95 % of testicular ligandin consisted of Yb and was used for functional studies as a Yb homodimer. Ya and Yb homodimer had glutathione S-transferase specific activities similar to those of native ligandin. Circular dichroism spectra showed that YaYa and YaYb had high affinity bilirubin binding sites but that YbYb had only low affinity binding sites. In the heterodimeric, Ya preferentially bound 35S-BSP covalently after incubation and ultraviolet irradiation. Although glutathione S-transferase activity was associated with each subunit of ligandin, the high affinity binding capacity of the protein was associated with Ya.
札幌医科大学の論文