Interaction between two domains of the P. yoelii MSP-1 protein detected using the yeast two-hybrid system.
Several model systems of plasmodia have demonstrated the potential of the merozoite surface protein, MSP-1, to induce protective immunity. However, little is known about the function of this protein or its interaction with other surface molecules that may also serve as immunological targets. To identify potentially significant inter- and intra-molecular interactions involving MSP-1, we have utilized the yeast two-hybrid system. A cDNA activation domain library was constructed from the erythrocytic stages of the murine malarial parasite Plasmodium yoelii yoelii 17XL. A 795 bp region of Py17XL MSP-1 (bait), homologous to the Plasmodium falciparum MSP1(33) fragment, was inserted into a Gal4p DNA binding domain vector and used to screen the activation domain library (target). Several randomly selected clones that demonstrated bait-target interaction were found to express overlapping regions of Py17XL MSP-1. Deletion constructs further localized the peptide fragments retaining interaction indicating that a region within the MSP-1(38) fragment interacts with the MSP-1 bait domain. Subsequent studies confirmed this interaction, as both peptides were co-precipitated from cell lysate by a peptide tag-specific antibody. It was observed that the interaction of these two fragments significantly increased the half-life of the MSP-1(38) within yeast cells. The specific interaction described here demonstrates the potential of this approach to elucidate additional inter- or intra-molecular interactions of Py17XL MSP1 and other malarial proteins.
Daly
T M
TM
Department of Microbiology and Immunology, MCP Hahnemann University, 2900 Queen Lane, Philadelphia, PA 19129, USA.
Long
C A
CA
Bergman
L W
LW
D000595:Amino Acid Sequence; D000818:Animals; D004912:Erythrocytes / Q000469:parasitology; D015723:Gene Library; D008288:Malaria / Q000469:parasitology; D008297:Male; D020066:Merozoite Surface Protein 1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010967:Plasmodium yoelii / Q000737:chemistry / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D017422:Sequence Analysis, DNA; D020798:Two-Hybrid System Techniques
D020066:Merozoite Surface Protein 1