Sanjay Ram

The contrasting mechanisms of serum resistance of Neisseria gonorrhoeae and group B Neisseria meningitidis.

Neisseria gonorrhoeae and Neisseria meningitidis have evolved intricate mechanisms to evade complement-mediated killing. Sialylation of gonococcal lipooligosaccharide (LOS) results in conversion of previously serum sensitive strains to unstable serum resistance, which is mediated by factor H binding. Porin (Por) is also instrumental in mediating stable serum resistance in gonococci. The 5th loop of certain gonococcal PorlAs binds factor H, which efficiently inactivates C3b to iC3b. Factor H glycan residues may be essential for factor H binding to certain Por1A strains. Por1A strains can also regulate the classical pathway by binding to C4b-binding protein (C4bp) probably via the 1st loop of the Por molecule. Certain serum resistant Por1 B strains can also regulate complement by binding C4bp through a loop other than loop 1. Purified C4b can inhibit binding of C4bp to Por 1B, but not Por1A, suggesting different binding sites on C4bp for the two Por types. Unlike serum resistant gonococci, resistant meningococci have abundant C3b on their surface, which is only partially processed to iC3b. The main mechanism of complement evasion by group B meningococci is inhibition of membrane attack complex (MAC) insertion by their polysaccharide capsule. LOS structure may act in concert with capsule to prevent MAC insertion. Meningococcal strains with Class 3 Por preferentially bind factor H, suggesting Class 3 Por acts as a receptor for factor H.

C4bp binding to porin mediates stable serum resistance of Neisseria gonorrhoeae

Screening of 29 strains of Neisseria gonorrhoeae revealed that 16/21 serum resistant strains and 0/8 serum sensitive strains bound C4bp, suggesting that C4bp binding to gonococci could contribute to serum resistance. C4bp bound to gonococci retained cofactor (C4b-degrading) function. Using allelic exchange to construct strains with hybrid Por1A/B molecules, we demonstrate that the N-terminal loop (loop 1) of Por1A is required for C4bp binding. Serum resistant Por1B gonococcal strains also bind C4bp via their Por molecule. Using allelic exchange and site-directed mutagenesis, we have shown that loops 5 and 7 together form a negatively charged C4bp binding domain. C4bp-Por1B interactions are ionic in nature (inhibited by high salt as well as by heparin), while the C4bp-Por1A bond is hydrophobic. mAbs directed against SCR1 of the alpha-chain of C4bp inhibit C4bp binding to both Por1A and Por1B. Furthermore, only recombinant C4bp mutant molecules that contain alpha-chain SCR1 bind both Por1A and Por1B gonococci, confirming that SCR1 contains Por binding sites. C4bp alpha-chain monomers do not bind strains with either Por molecule, suggesting that the polymeric form of C4bp is required for binding to gonococci. Inhibition of C4bp binding to serum resistant Por1A and Por1B strains in a serum bactericidal assay using fAb fragments against C4bp SCR1 results in complete killing at 30 min of otherwise fully serum resistant strains in only 10% normal serum, underscoring the role of C4bp in mediating gonococcal serum resistance.