Human
C-reactive protein (CRP) protects mice from lethal
Streptococcus pneumoniae infection when injected into mice within the range of 6 h before to 2 h after the administration of pneumococci. Because CRP binds to
phosphocholine-containing substances and subsequently activates the
complement system, it has been proposed that the antipneumococcal function of CRP requires the binding of CRP to
phosphocholine moieties present in pneumococcal cell wall C-
polysaccharide. To test this proposal experimentally, in this study, we utilized a new CRP mutant incapable of binding to
phosphocholine. Based on the structure of CRP-
phosphocholine complexes, which showed that Phe(66), Thr(76), and Glu(81) formed the
phosphocholine-binding pocket, we constructed a CRP mutant F66A/T76Y/E81A in which the pocket was blocked by substituting Tyr for Thr(76). When compared with wild-type CRP, mutant CRP bound more avidly to
phosphoethanolamine and could be purified by affinity chromatography using
phosphoethanolamine-conjugated
Sepharose. Mutant CRP did not bind to
phosphocholine, C-
polysaccharide, or pneumococci. Mutant CRP was free in the mouse serum, and its rate of clearance in vivo was not faster than that of wild-type CRP. When either 25 μg or 150 μg of CRP was administered into mice, unlike wild-type CRP, mutant CRP did not protect mice from lethal
pneumococcal infection. Mice injected with mutant CRP had higher mortality rates than mice that received wild-type CRP. Decreased survival was due to the increased
bacteremia in mice treated with mutant CRP. We conclude that the
phosphocholine-binding pocket on CRP is necessary for CRP-mediated initial protection of mice against lethal
pneumococcal infection.