Identification of specific Hep G2 cell binding regions in Plasmodium falciparum sporozoite-threonine-asparagine-rich protein (STARP)

It has been demonstrated that Plasmodium falciparum sporozoite threonine-asparagine-rich protein (PfSTARP) is located on the sporozoite surface. This protein's non-overlapping consecutive peptides were synthesised and tested in Hep G2 cell binding assays. Twelve high activity binding peptides (HABPs) were identified in the resulting 31 peptides. Three were found in 5′ non-repeat region (amino acids 41-80). Peptides 20546 (⁴¹VIKHNRFLSEYQSNFLGGGY⁶⁰), 20547 (⁶¹SAALKLVNSKKSGTNVNVTKY⁸⁰) and 20548 (⁸¹NSENTNTNNNIPESSSTYTN¹⁰⁰) were located in the conserved amino terminal region, as well as peptide 20548 which shared the sequence with the M region (amino acids 85-134). Six HABPs were located in region 10 (Rp10) (STDNNNTKTI). HABPs 20569 (⁵⁰¹TSDDELNKDSCDYSEEKENI⁵²⁰) and 20570 (⁵²¹KSMINAYLDKLDLETVRKIH⁴⁰) were found in 3′ non-repeat region. All these HABPs showed saturable binding and presented dissociation constants between 18 and 219nM. The number of binding sites per Hep G2 cell ranged from 45,000 to 370,000. High binding peptides' critical amino acids involved in Hep G2 cell binding were determined by competition binding assays. SDS-PAGE results showed that both peptides 20570 and 20547 had at least two different sets of 44 and 38kDa HABP receptors on Hep G2 cells. Specific modification of peptide 20546 and 20570 critical binding residues rendered these peptides immunogenic in Aotus monkeys, inducing high antibody titres against sporozoites, as assessed by IFA.