Molecular and biological characterization of a mannan-binding lectin from the holothurian Apostichopus japonicus.
To elucidate the origin and evolution of mannan-binding lectins (MBL), a new C-type lectin (CTL) specific for high-mannose glycans (MBL-AJ) was isolated from the coelomic plasma of the holothurian Apostichopus japonicus. MBL-AJ has oligomeric forms with identical 17-kDa subunits on SDS-PAGE. Among natural ligands, lectin hemagglutination activity was competitively inhibited by extracellular low-branched, but not high-branched, alpha-D-mannans isolated from marine halophilic bacteria and composed of alpha-1,2 and alpha-1,6 linked D-mannose residues. This suggests that the lectin interacts with backbone or inner side chain mannose residues, but not with terminal ones. The activity of the lectin was Ca(2+)-, pH-, and temperature-dependent. MBL-AJ cDNA was cloned from a holothurian coelomocyte cDNA library. The subunit of the mature protein has 159 amino acids and a single carbohydrate-recognition domain (CRD) of CTL. CRD contains a Glu-Pro-Asp amino acid sequence (EPN-motif) conserved for all known MBLs. A monospecific polyclonal antibody against MBL-AJ was obtained using the 34-kDa lectin dimer as an immunogen. The MBL-AJ has demonstrated immunochemical identity to the earlier isolated mannan-binding CTL from another holothurian, Cucumaria japonica. But a more interesting finding was cross-reactivity of MBL-AJ and human serum MBL detected by the antibody against MBL-AJ. Taking into consideration such MBL-AJ peculiarities as its carbohydrate specificity, the presence of a conserved region forming the mannose-binding site, common antigenic determinants with human MBL, and participation in defense reactions, it is possible that MBL-AJ belongs to the family of evolutionary conserved mannan-binding proteins.
Bulgakov
Aleksandr A
AA
Pacific Institute of Bioorganic Chemistry Far Eastern Branch, Russian Academy of Science, Vladivostok, Russia. aabulgakov@yahmo.com
Eliseikina
Marina G
MG
Petrova
Irina Yu
IY
Nazarenko
Evgeny L
EL
Kovalchuk
Svetlana N
SN
Kozhemyako
Valery B
VB
Rasskazov
Valery A
VA
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002241:Carbohydrates / Q000737:chemistry; D019281:Dimerization; D005976:Glutaral / Q000737:chemistry; D006388:Hemagglutinins / Q000737:chemistry; D006801:Humans; D037102:Lectins / Q000737:chemistry*; D008024:Ligands; D008351:Mannans / Q000737:chemistry*; D037601:Mannose-Binding Lectin / Q000737:chemistry; D008954:Models, Biological; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D047528:Stichopus / Q000378:metabolism*
D002241:Carbohydrates; D006388:Hemagglutinins; D037102:Lectins; D008024:Ligands; D008351:Mannans; D037601:Mannose-Binding Lectin; D005976:Glutaral