IDENTIFICATION AND HETEROLOGOUS EXPRESSION OF A NEW DENSE GRANULE PROTEIN (GRA7) FROM TOXOPLASMA-GONDII

Immunoscreening of an expression library constructed with Toxoplasma g ondii tachyzoite mRNA with sera from toxoplasmosis-positive humans has led to the identification of a new parasite antigen. Sequence analysi s of the gene encoding this antigen allowed the calculation of the the oretical molecular mass (25 857 Da) and showed that the protein contai ns a putative signal sequence. The C-terminal region contains two hydr ophobic regions, the last of which has the characteristics of a membra ne-spanning domain. When the protein was heterologously expressed in E . coil and tested by Western blot, it reacted with the human sera orig inally used for screening. The new antigen also reacted with a monoclo nal antibody raised against the entire parasite. Ultrastructural analy sis showed that the protein is localized in the dense granules. After host cell invasion, the protein is secreted into the vacuolar network, the parasitophorous vacuole membrane, and into extensions protruding in the cytoplasm. Therefore, it is suggested to designate this new den se granule protein GRA7, following the established nomenclature for th is protein family. (C) 1998 Elsevier Science B.V. All rights reserved.