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J. Biol. Chem., Vol. 282, Issue 48, 34877-34887, November 30, 2007

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Cryptosporidium p30, a Galactose/N-Acetylgalactosamine-specific Lectin, Mediates Infection in Vitro^*

Najma Bhat, Angela Joe¹, Mercio PereiraPerrin, and Honorine D. Ward²

From the Division of Geographic Medicine and Infectious Diseases, Tufts-New England Medical Center and the Department of Pathology, Tufts University School of Medicine, Boston, Massachusetts 02111

Cryptosporidium sp. cause human and animal diarrheal disease worldwide. The molecular mechanisms underlying Cryptosporidium attachment to, and invasion of, host cells are poorly understood. Previously, we described a surface-associated Gal/GalNAc-specific lectin activity in sporozoites of Cryptosporidium parvum. Here we describe p30, a 30-kDa Gal/GalNAc-specific lectin isolated from C. parvum and Cryptosporidium hominis sporozoites by Gal-affinity chromatography. p30 is encoded by a single copy gene containing a 906-bp open reading frame, the deduced amino acid sequence of which predicts a 302-amino acid, 31.8-kDa protein with a 22-amino acid N-terminal signal sequence. The p30 gene is expressed at 24–72 h after infection of intestinal epithelial cells. Antisera to recombinant p30 expressed in Escherichia coli react with an 30-kDa protein in C. parvum and C. hominis. p30 is localized to the apical region of sporozoites and is predominantly intracellular in both sporozoites and intracellular stages of the parasite. p30 associates with gp900 and gp40, Gal/GalNAc-containing mucin-like glycoproteins that are also implicated in mediating infection. Native and recombinant p30 bind to Caco-2A cells in a dose-dependent, saturable, and Gal-inhibitable manner. Recombinant p30 inhibits C. parvum attachment to and infection of Caco-2A cells, whereas antisera to the recombinant protein also inhibit infection. Taken together, these findings suggest that p30 mediates C. parvum infection in vitro and raise the possibility that this protein may serve as a target for intervention.

Received for publication, August 20, 2007 , and in revised form, September 28, 2007.

The nucleotide sequence(s) reported in this paper has been submitted to the GenBank^TM/EBI Data Bank with accession number(s) AY308041 [GenBank] and AY308040 [GenBank] .

^* This work was supported by United States Public Health Service Grants AI45194, AI52786 (to H. W.), and NS40574 (to M. P.) and by the Center for Gastroenterology Research on Absorptive and Secretory Processes, Digestive Disease Center, United States Public Health Service Grant P30 DK34928. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ Present address: Dept. of Microbiology and Immunology, University of Melbourne, 3010 Parkville, Australia.

² To whom correspondence should be addressed: Division of Geographic Medicine and Infectious Diseases, Tufts-New England Medical Center, Box 041, 750 Washington St., Boston, MA 02111. Tel.: 617-636-7022; Fax: 617-636-5292; E-mail: hward{at}tufts-nemc.org.

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