Abstract :
[en] A sequence-based prediction method was employed to identify three ligand-binding
domains in transferrin-binding protein B (TbpB) of Neisseria meningitidis strain
B16B6. Site-directed mutagenesis of residues located in these domains has led to
the identification of two domains, amino acids 53 to 57 and 240 to 245, which are
involved in binding to human transferrin (htf). These two domains are conserved
in an alignment of different TbpB sequences from N. meningitidis and Neisseria
gonorrhoeae, indicating a general functional role of the domains. Western blot
analysis and BIAcore and isothermal titration calorimetry experiments
demonstrated that site-directed mutations in both binding domains led to a
decrease or abolition of htf binding. Analysis of mutated proteins by circular
dichroism did not provide any evidence for structural alterations due to the
amino acid replacements. The TbpB mutant R243N was devoid of any htf-binding
activity, and antibodies elicited by the mutant showed strong bactericidal
activity against the homologous strain, as well as against several heterologous
tbpB isotype I strains.
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