Crystal structure of native and Cd/Cd-substituted Dioclea guianensis seed lectin. A novel manganese-binding site and structural basis of dimer-tetramer association.
Diocleinae legume lectins are a group of oligomeric proteins whose subunits display a high degree of primary structure and tertiary fold conservation but exhibit considerable diversity in their oligomerisation modes. To elucidate the structural determinants underlaying Diocleinae lectin oligomerisation, we have determined the crystal structures of native and cadmium-substituted Dioclea guianensis (Dguia) seed lectin. These structures have been solved by molecular replacement using concanavalin (ConA) coordinates as the starting model, and refined against data to 2.0 A resolution. In the native (Mn/Ca-Dguia) crystal form (P4(3)2(1)2), the asymmetric unit contains two monomers arranged into a canonical legume lectin dimer, and the tetramer is formed with a symmetry-related dimer. In the Cd/Cd-substituted form (I4(1)22), the asymmetric unit is occupied by a monomer. In both crystal forms, the tetrameric association is achieved by the corresponding symmetry operators. Like other legume lectins, native D. guianensis lectin contains manganese and calcium ions bound in the vicinity of the saccharide-combining site. The architecture of these metal-binding sites (S1 and S2) changed only slightly in the cadmium/cadmium-substituted form. A highly ordered calcium (native lectin) or cadmium (Cd/Cd-substituted lectin) ion is coordinated at the interface between dimers that are not tetrameric partners in a similar manner as the previously identified Cd(2+) in site S3 of a Cd/Ca-ConA. An additional Mn(2+) coordination site (called S5), whose presence has not been reported in crystal structures of any other homologous lectin, is present in both, the Mn/Ca and the Cd/Cd-substituted D. guianensis lectin forms. On the other hand, comparison of the primary and quaternary crystal structures of seed lectins from D. guianensis and Dioclea grandiflora (1DGL) indicates that the loop comprising residues 117-123 is ordered to make interdimer contacts in the D. grandiflora lectin structure, while this loop is disordered in the D. guianensis lectin structure. A single amino acid difference at position 131 (histidine in D. grandiflora and asparagine in D. guianensis) drastically reduces interdimer contacts, accounting for the disordered loop. Further, this amino acid change yields a conformation that may explain why a pH-dependent dimer-tetramer equilibrium exists for the D. guianensis lectin but not for the D. grandiflora lectin.
Wah
D A
DA
Centro de Investigaciones Biológicas, C.S.I.C., Madrid, Spain.
Romero
A
A
Gallego del Sol
F
F
Cavada
B S
BS
Ramos
M V
MV
Grangeiro
T B
TB
Sampaio
A H
AH
Calvete
J J
JJ
D001665:Binding Sites; D002104:Cadmium / Q000378:metabolism*; D018360:Crystallography, X-Ray; D019281:Dimerization; D006863:Hydrogen-Ion Concentration; D037102:Lectins / Q000737:chemistry* / Q000378:metabolism*; D019684:Magnoliopsida / Q000737:chemistry*; D008345:Manganese / Q000378:metabolism*; D008958:Models, Molecular; D037121:Plant Lectins; D010940:Plant Proteins / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D012639:Seeds / Q000737:chemistry; D013816:Thermodynamics
D037102:Lectins; D037121:Plant Lectins; D010940:Plant Proteins; D002104:Cadmium; D008345:Manganese