Journal of’ General Microbiology (1992), 138, 2663-2672. Printed in Great Britain 2663 Purification and characterization of a.surface lectin from the nematode-trapping fungus Arthrobotrys ofigospora STEFAN RosEN,’ * €30 EK,~ LARS RASK~ and ANDERS TUNLID’ ,* ’ Department of’ Microbial Ecology, Lund University, Ecology Building, Helgonavagen 5, S-223 62 Lund, Sweden Department of’ Chemical Ecology and Ecotoxicology, Lund University, Ecology Building, Helgonavagen 5, S-223 62 Lund, Sweden Department of’ Cell Research, Swedish University of Agricultural Science, Box 7055, S-750 07 Uppsala, Sweden (Received 8 June 1992; revised I0 August 1992; accepted 18 August 1992) Several studies have indicated that the capture of nematodes by the nematophagous fungus Arthrobotrys oligospora is mediated by a lectin on the fungal surface. One of the major surface proteins of this fungus showed haemagglutinating activity and was isolated by affinity chromatography using a mucin Sepharose column. Biochemical analysis showed that the protein was a dimeric glycoprotein with a molecular mass of 36 kDa and an isoelectric point of pH 6.5, and contained no sulphur amino acids. The protein was N-terminally blocked; four internal peptides were sequenced, and showed no significant similarity to sequences in the Swiss-Prot or PIR databases. The haemagglutinating activity of the isolated protein was not inhibited by any of the mono- or disaccharides tested, but it was inhibited by the glycoproteins fetuin and m u c h The haemagglutinating activity changed after incubating the protein in buffers of different pH, with maximal activity at pH 11.0 and no activity at pH 2.8. The lectin was tested for different enzymic activities but none were detected. Analysis of the haemagglutinating activity in various cell fractions indicated that the protein was associated with extracellular polymer layers and with the cell wall of the fungus. About the same amount of the haemagglutinating protein was recovered from samples of vegetative mycelium and of mycelium containing nematode-trapping cells. Introduction Fungal adhesion to host surfaces is an important event in the colonization of humans, animals and plants by fungi, and subsequent pathogenesis. Knowledge of fungal adhesion and the factors that influence fungal attach- ment is still, however, limited (Kennedy, 1990), and no fungal adhesive compound has been fully characterized (Nicholson & Epstein, 1991). Carbohydrate-protein (lectin) interactions have been proposed to mediate adhesion in several fungal-host systems, plant patho- gens, fungal-fungal interactions (mycoparasitism), fun- gal-algal interactions and nematophagous fungi (Nordbring-Hertz & Chet, 1986; Manocha & Chen, 1989). One of the first systems to suggest lectin-mediated adhesion was the nematophagus fungus Arthrobotrys oligospora. This fungus captures nematodes using adhe- sive polymers present on special hyphae (traps) forming * Author for correspondence. Tel. 46 46 10 96 14; fax 46 46 10 41 58. 0001-7624 O 1992 SGM a three-dimensional network (Barron, 1977 ; Nordbring- Hertz, 1988). Flooding a colony of A. oligospora with a 20 mM-N-aCetyl-galaCtOSamine (GalNac) solution inhib- its capture, while treatment of the nematodes with this carbohydrate does not inhibit adhesion (Nordbring- Hertz & Mattiasson, 1979). Small amounts of a GalNac- binding protein have been purified on a GalNac-affinity column (Borrebaeck et al., 1984; Premachandran & Pramer, 1984), and it has been found only in trap- containing mycelium and not in vegetative mycelium (Borrebaeck et al., 1984, 1985). Using a soybean agglutinin (SBA ; specificity for GalNac) affinity gel, Borrebaeck et a!. (1985) isolated an apparent GalNac specific receptor protein with a molecular mass of approx. 65 kDa from the cuticle of the nematode. Involvement of lectin-mediated adhesion was further confirmed by demonstrating that entrapment was re- duced when the nematode was treated with a protein purified on a GalNac column (Premachandran & Pramer, 1984). Recent studies of the adhesion mechanism in A.