Characterization and use of biotinylated Escherichia coli K99 lectin.

K99 lectin from Escherichia coli was purified and biotinylated via the amino groups of lysine residues using N-biotinyl-6-amino-caproic acid N-hydroxysuccinimide ester (BcapNHS). Biotin was detected on Lys-47 and Lys-87. It was previously demonstrated (Jacobs, A.A.C., Van den Berg, P.A., Bak, H.J. and De Graaf, F.K. (1986) Biochim. Biophys. Acta 872, 92-97) that modification of lysine residues 132 and 133 with 4-chloro-3,5-dinitrobenzoate (CDNB) resulted in the loss of the binding capacity of K99 fimbriae. Due to the higher size of the biotin derivative compared to CDNB, Lys-132 or Lys-133, essential for the biological activity, were not modified. The biotinylation did not cause the loss of the haemagglutinating activity but was sufficient to permit detection of the lectin by streptavidin. A flow cytometric analysis was used for the detection of the receptors on the surface of erythrocytes.