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 go urami, Trichogaster trichopterus, with the use of (NH4)(2)SO4 fractionation , affinity chromatography and gel filtration chromatography. Electrophoreti c analyses and mass spectrometric study of purified BGL showed that the lec tin is composed of two isoforms with native molecular masses estimated to b e 65 and 66 kDa, and two subunits of 32 and 34 kDa on SDS-PAGE under nonred ucing conditions. Upon reduction with 20 mM dithiothreitol (DTT), BGL showe d 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 sha re the same sequence (HGEENRXGPR) and show no significant homology with any known proteins. The BGL agglutinating activity is specifically inhibited b y N-acetyl-D-galactosamine and N-acetyl-D-glucosamine, and to a lesser degr ee by D-(+)-mannose, but not by D-(+)-galactose, D-(+)-glucose, maltose or N-acetyl-D-mannosamine. Haemagglutination assay showed that BGL is more spe cific for rabbit than mouse, chicken, rat or guinea pig erythrocytes, and h aemagglutination was Ca2+-dependent. In addition, BGL could agglutinate a r ange of micro-organisms and yeast cells, with the exception of some fish pa thogens, such as Aeromonas hydrophila (strains: PPD 134/91 and PPD 11/90) a nd Vibrio harveyi (strain: W618). Localisation of BGL by fluorescein isothi ocyanate (FITC)-labelled antibodies revealed that the lectin is associated with the cell surface of fish leukocytes. (C) 2000 Academic Press.