The structure of a tunicate C-type lectin from Polyandrocarpa misakiensis complexed with D-galactose

C-type lectins are calcium-dependent carbohydrate-recognising proteins. Iso thermal titration calorimetry of the C-type Polyandrocarpa lectin (TC14) fr om the tunicate Polyandrocarpa misakiensis revealed the presence of a singl e calcium atom per monomer with a dissociation constant of 2.6 mu M, and co nfirmed the specificity of TC14 for D-galactose and related monosaccharides . We have determined the 2.2 Angstrom X-ray crystal structure of Polyandroc arpa lectin complexed with D-galactose. Analytical ultracentrifugation reve aled that TC14 behaves as a dimer in solution. This is reflected by the pre sence of two molecules in the asymmetric unit with the dimeric interface fo rmed by antiparallel pairing of the two N-terminal beta-strands and hydroph obic interactions. TC14 adopts a typical C-type lectin fold with difference s in structure from other C-type lectins mainly in the diverse loop regions and in the second alpha-helix, which is involved in the formation of the d imeric interface. The D-galactose is bound through coordination of the 3 an d 4-hydroxyl oxygen atoms with a bound calcium atom. Additional hydrogen bo nds are formed directly between serine, aspartate and glutamate side-chains of the protein and the sugar 3 and 4-hydroxyl groups. Comparison of the ga lactose binding by TC14 with the mannose binding by rat mannose-binding pro tein reveals how monosaccharide specificity is achieved in this lectin. A t ryptophan side-chain close to the binding site and the distribution of hydr ogen-bond accepters and donors around the 3 and 4-hydroxyl groups of the su gar are essential determinants of specificity. These elements are, however, arranged in a very different way than in an engineered galactose-specific mutant of MBPA. Possible biological functions can more easily be understood from the fact that TC14 is a dimer under physiological conditions. (C) 199 9 Academic Press.