Isolation of mannose-binding C-type lectin from sea lamprey ( Petromyzon marinus) plasma and binding to Aeromonas salmonicida

Abstract

The sea lamprey ( Petromyzon marinus) is a parasitic cartilaginous fish of the North American Great Lakes and a predator of many bony fish species of commercial importance to the fishing industry. Mannose-binding C-type lectin (MBL) was isolated by mannan–agarose affinity chromatography from sea lamprey plasma. Mannose-binding lectin has not before been identified and quantitated in the plasma of this sea lamprey species. The affinity-purified and 2-ME reduced lamprey MBL showed two bands of 35 kDa and 65 kDa by SDS-PAGE and Western blotting using guinea pig anti-MBL IgG as the primary antibody. Amino acid composition analysis (mol%) of the purified lamprey MBL found high amounts of histidine, threonine, tyrosine and phenylalanine present when compared with three other vertebrate MBLs. N-terminal amino acid sequencing by Edman degradation for the first 10 residues gave XXXTKGCPDA. Lamprey plasma contained 261 μg of MBL/ml of plasma. Plasma protein concentration was 40.1 mg/ml. Lamprey MBL was present then in plasma at 6.5 μg MBL/mg total protein. The sea lamprey MBL also specifically binds to mannose on the surface of the pathogen Aeromonas salmonicida. The presence of MBL in high concentration in lamprey plasma could be important in their innate immunity and resistance to infection. This study describes the presence of MBL in sea lamprey plasma and evidence for a C-type lectin complement pathway of innate immunity.