TY - JOUR
T1 - Using mutagenesis and structural biology to map the binding site for the plasmodium falciparum merozoite protein PfRh4 on the human immune adherence receptor
AU - Park, Hyon Ju
AU - Guariento, Mara
AU - Maciejewski, Mateusz
AU - Hauhart, Richard
AU - Tham, Wai Hong
AU - Cowman, Alan F.
AU - Schmidt, Christoph Q.
AU - Mertens, Haydyn D.T.
AU - Kathryn Liszewski, M.
AU - Hourcade, Dennis E.
AU - Barlow, Paul N.
AU - Atkinson, John P.
PY - 2014/1/3
Y1 - 2014/1/3
N2 - To survive and replicate within the human host, malaria parasites must invade erythrocytes. Invasion can be mediated by the P. falciparum reticulocyte-binding homologue protein 4 (PfRh4) on the merozoite surface interacting with complement receptor type 1 (CR1, CD35) on the erythrocyte membrane. The PfRh4 attachment site lies within the three N-terminal complement control protein modules (CCPs 1-3) of CR1, which intriguingly also accommodate binding and regulatory sites for the key complement activation-specific proteolytic products, C3b and C4b. One of these regulatory activities is decay-accelerating activity. Although PfRh4 does not impact C3b/C4b binding, it does inhibit this convertase disassociating capability. Here, we have employed ELISA, co-immunoprecipitation, and surface plasmon resonance to demonstrate that CCP 1 contains all the critical residues for PfRh4 interaction. We fine mapped by homologous substitution mutagenesis the PfRh4-binding site on CCP 1 and visualized it with a solution structure of CCPs 1-3 derived by NMR and small angle x-ray scattering. We crossvalidated these results by creating an artificial PfRh4-binding site through substitution of putative PfRh4-interacting residues from CCP 1 into their homologous positions within CCP 8; strikingly, this engineered binding site had an ∼30-fold higher affinity for PfRh4 than the native one in CCP 1. These experiments define a candidate site on CR1 by which P. falciparum merozoites gain access to human erythrocytes in a non-sialic acid-dependent pathway of merozoite invasion.
AB - To survive and replicate within the human host, malaria parasites must invade erythrocytes. Invasion can be mediated by the P. falciparum reticulocyte-binding homologue protein 4 (PfRh4) on the merozoite surface interacting with complement receptor type 1 (CR1, CD35) on the erythrocyte membrane. The PfRh4 attachment site lies within the three N-terminal complement control protein modules (CCPs 1-3) of CR1, which intriguingly also accommodate binding and regulatory sites for the key complement activation-specific proteolytic products, C3b and C4b. One of these regulatory activities is decay-accelerating activity. Although PfRh4 does not impact C3b/C4b binding, it does inhibit this convertase disassociating capability. Here, we have employed ELISA, co-immunoprecipitation, and surface plasmon resonance to demonstrate that CCP 1 contains all the critical residues for PfRh4 interaction. We fine mapped by homologous substitution mutagenesis the PfRh4-binding site on CCP 1 and visualized it with a solution structure of CCPs 1-3 derived by NMR and small angle x-ray scattering. We crossvalidated these results by creating an artificial PfRh4-binding site through substitution of putative PfRh4-interacting residues from CCP 1 into their homologous positions within CCP 8; strikingly, this engineered binding site had an ∼30-fold higher affinity for PfRh4 than the native one in CCP 1. These experiments define a candidate site on CR1 by which P. falciparum merozoites gain access to human erythrocytes in a non-sialic acid-dependent pathway of merozoite invasion.
UR - http://www.scopus.com/inward/record.url?scp=84891707973&partnerID=8YFLogxK
U2 - 10.1074/jbc.M113.520346
DO - 10.1074/jbc.M113.520346
M3 - Article
C2 - 24214979
AN - SCOPUS:84891707973
SN - 0021-9258
VL - 289
SP - 450
EP - 463
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 1
ER -