Galactosyl-binding Lectins from the Tunicate Didernnurn candidurn

acid sequence of two galactosyl-binding lectins D. candidum lectins I and I1 (DCL-I and DCL-11) from the plasma of this protochordate species. Both lectins were purified by affinity chromatography (on acid-treated Sepharose 4B and asialofetuin conjugated to Sepharose 4B) to homogeneity as judged by immunoelectrophoresis, size exclusion chromatography on high performance liquid chromatography, and polyacrylamide gel electrophoresis. Isoelectric focusing in polyacrylamide gels revealed that DCL-I focuses as a family of bands at pH 3.8-5.2, while DCL-I1 focuses at pH 9.2-10.2. Gas chromatography analyses of alditol acetate derivatives indicated that no carbohydrate components are associated with the lectins. Approximate subunit molecular weights estimated by polyacrylamide gel electrophoresis and size exclusion chromatography on high performance liquid chromatography in 6 M guanidine HCl under reducing conditions were 13,400-14,500 for DCL-I and 14,500-15,500 for DCL-11. Native molecular weights estimated by sedimentation equilibrium were 56,600 (DCL-I) and 57,500 (DCL-11), indicating that both species are constituted by four equal-sized subunits. Frictional ratios suggested that both lectins are globular proteins. Using rabbit antisera, the two molecules appeared serologically distinct. The extinction coefficient for DCL-I was = 2.62 ml mg” cm”. Circular dichroism analyses of DCL-I suggested 29% a-helix and 37% B-structure in the protein. Excitation/emission fluorescence spectra for DCL-I yielded maximum excitation and emission wavelengths at 288 and 330 nm, respectively. Amino acid compositions of DCL-I and DCL-I1 differed mainly in the proportions of aspartic and glutamic acids, serine, alanine, cysteine, valine, phenylalanine, and histidine. Amino acid compositions of DCL-I and DCL-I1 were compared to each other and to immunoglobulins and putative recognition molecules by the parameter SAQ. DCL-I exhibited similarities in amino acid composition to lectins from the tunicate Halocynthia pyriformis, the lamprey Petromyzon marinus,