Isolation and characterisation of a serum lectin from blue gourami, Trichogaster trichopterus (Pallus).
A novel lectin, designated BGL, has been purified from the serum of blue gourami, Trichogaster trichopterus, with the use of (NH4)2SO4 fractionation, affinity chromatography and gel filtration chromatography. Electrophoretic analyses and mass spectrometric study of purified BGL showed that the lectin is composed of two isoforms with native molecular masses estimated to be 65 and 66 kDa, and two subunits of 32 and 34 kDa on SDS-PAGE under non-reducing conditions. Upon reduction with 20 mM dithiothreitol (DTT), BGL showed two close bands of 27 and 29 kDa. After isoelectric focusing, the lectin focused as close double bands at pH 5.6. The N-termini of both isoforms share the same sequence (HGEENRXGPR) and show no significant homology with any known proteins. The BGL agglutinating activity is specifically inhibited by N-acetyl-D-galactosamine and N-acetyl-D-glucosamine, and to a lesser degree by D-(+)-mannose, but not by D-(+)-galactose, D-(+)-glucose, maltose or N-acetyl-D-mannosamine. Haemagglutination assay showed that BGL is more specific for rabbit than mouse, chicken, rat or guinea pig erythrocytes, and haemagglutination was Ca2+-dependent. In addition, BGL could agglutinate a range of micro-organisms and yeast cells, with the exception of some fish pathogens, such as Aeromonas hydrophila (strains: PPD 134/91 and PPD 11/90) and Vibrio harveyi (strain: W618). Localisation of BGL by fluorescein isothiocyanate (FITC)-labelled antibodies revealed that the lectin is associated with the cell surface of fish leukocytes.
Fock
W L
WL
Department of Biological Sciences, National University of Singapore, Singapore.
Chen
C L
CL
Lam
T J
TJ
Sin
Y M
YM
D000371:Agglutination; D000818:Animals; D002241:Carbohydrates / Q000737:chemistry; D002847:Chromatography, Agarose / Q000662:veterinary; D002850:Chromatography, Gel / Q000662:veterinary; D004591:Electrophoresis, Polyacrylamide Gel / Q000662:veterinary; D006384:Hemagglutination; D005779:Immunodiffusion / Q000662:veterinary; D007525:Isoelectric Focusing / Q000662:veterinary; D037102:Lectins / Q000737:chemistry / Q000302:isolation & purification*; D008858:Microscopy, Phase-Contrast / Q000662:veterinary; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010473:Perciformes / Q000502:physiology*; D010586:Phagocytes / Q000737:chemistry; D020539:Sequence Analysis, Protein; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000662:veterinary
D002241:Carbohydrates; D037102:Lectins