Isolation and expression of the gene for a major surface protein of Giardia lamblia.

Abstract

To study the interactions between the parasitic protozoan Giardia lamblia and its environment, we have cloned the gene that encodes the two major surface-labeled trophozoite protein species. Sequence analysis of this gene reveals a single open reading frame specifying a hydrophilic, cysteine-rich (11.8%) protein of 72.5-kDa molecular mass with an amino-terminal signal peptide and a postulated hydrophobic membrane-spanning anchor region near the carboxyl terminus. Most of the cysteine residues (58 of 84) are in the motif Cys-Xaa-Xaa-Cys, which is dispersed 29 times throughout the sequence. Antibodies against the recombinant protein react with the entire surface of live trophozoites, including flagella and adhesive disc. These antibodies inhibit trophozoite attachment, prevent growth, and immunoprecipitate the major =66-and 85-kDa proteins from surface-labeled live trophozoites. The recombinant Escherichia coli also expresses polypeptides of -66-and 85-kDa molecular mass, which are not fusion proteins. This suggests that the processing and/or conformational changes that lead to production of these two peptide species in E. coli reflect those that occur in Giardia. The abundance of cysteine residues suggests that the native proteins on the parasite surface may contain numerous disulfide bonds, which would promote resistance to intestinal fluid proteases and to the detergent activity of bile salts and would help to explain the survival of Giardia in the human small intestine.

Distinct genetic groups of Giardia intestinalis distinguished by restriction fragment length polymorphisms

“…and amino acid level, with the tsa417 gene (Gillin et al, 1990) that encodes a M , 72500 surface antigen on trophozoites of G. intestinalis isolate WB.…”

“…These were prepared from two genomic DNA fragments (from the Ad-1 G. intestinalis isolate) that had been cloned by ligation into the BarnHI cloning site of plasmid pUC8. Probes lA, 1B and 1C were derived from the recombinant plasmid pKKl1, which contains a gene, tspl I, that encodes a predicted M, 69000 cysteine-rich trophozoite surface protein, TSPll , related to the TSA417 antigen described by Gillin et al (1990). Probe 2 was prepared from an unrelated genomic fragment isolated from plasmid pKK40 (K. Khanna, P. Ey & G. Mayrhofer, unpublished results).…”

Characterisation of the subtelomeric regions of Giardia lamblia genome isolate WBC6

“…The genome isolate, WB clone C6 (ATCC #50803), was the major isolate used for this study (McArthur et al, 2000;Gillin et al, 1990). Trophozoites were grown in modified TYI-S-33 medium (Keister, 1983).…”

Lectin blot studies on proteins of Myxobolus cerebralis, the causative agent of whirling disease

“…To counter the host response, protozoans rely on antigen mimicry (Damian 1987, Inal 2004) and antigen variation (Borst et al 1996, Rudenko et al 1998, processes which involve surface glycoproteins such as glycoepitopes of N-glycosides (Burghaus et al 1999, Yang et al 1999 and O-glycosides (Dieckmann-Schuppert et al 1993, Khan et al 1997. In Trypanosoma brucei (Borst et al 1996, Ferguson 1997, 1999 and Giardia lamblia (Gillin et al 1990, Gillin & Reiner 1996, Nash 1992), a variant surface glycoprotein (VSG) is expressed sequentially to evade the host's immune reaction.…”