Characterization of the fetuin-binding fraction of Neospora caninum tachyzoites and its potential involvement in host-parasite interactions.
Terminal sialic acid residues on surface-associated glycoconjugates mediate host cell interactions of many pathogens. Addition of sialic acid-rich fetuin enhanced, and the presence of the sialidiase inhibitor 2-deoxy-2,3-dehydro-N-acetylneuraminic acid reduced, the physical interaction of Neospora caninum tachyzoites and bradyzoites with Vero cell monolayers. Thus, Neospora extracts were subjected to fetuin-agarose affinity chromatography in order to isolate components potentially interacting with sialic acid residues. SDS-PAGE and silver staining of the fetuin binding fraction revealed the presence of a single protein band of approximately 65 kDa, subsequently named NcFBP (Neospora caninum fetuin-binding protein), which was localized at the apical tip of the tachyzoites and was continuously released into the surrounding medium in a temperature-independent manner. NcFBP readily interacted with Vero cells and bound to chondroitin sulfate A and C, and anti-NcFBP antibodies interfered in tachyzoite adhesion to host cell monolayers. In additon, analysis of the fetuin binding fraction by gelatin substrate zymography was performed, and demonstrated the presence of two bands of 96 and 140 kDa exhibiting metalloprotease-activity. The metalloprotease activity readily degraded glycosylated proteins such as fetuin and bovine immunoglobulin G heavy chain, whereas non-glycosylated proteins such as bovine serum albumin and immunoglobulin G light chain were not affected. These findings suggest that the fetuin-binding fraction of Neospora caninum tachyzoites contains components that could be potentially involved in host-parasite interactions.
Vonlaufen
N
N
Institute of Parasitology, Vetsuisse Faculty, University of Berne, Länggass-Strasse 122, CH-3012 Berne, Switzerland.
Naguleswaran
A
A
Gianinazzi
C
C
Hemphill
A
A
D000818:Animals; D000913:Antibodies, Protozoan / Q000032:analysis / Q000378:metabolism; D002448:Cell Adhesion; D002522:Chlorocebus aethiops; D006790:Host-Parasite Interactions / Q000502:physiology*; D008018:Life Cycle Stages; D045726:Metalloproteases / Q000378:metabolism; D019158:N-Acetylneuraminic Acid / Q000378:metabolism; D018688:Neospora / Q000201:enzymology / Q000378:metabolism*; D015800:Protozoan Proteins / Q000378:metabolism*; D013696:Temperature; D014709:Vero Cells; D000509:alpha-Fetoproteins / Q000378:metabolism*
D000913:Antibodies, Protozoan; D015800:Protozoan Proteins; D000509:alpha-Fetoproteins; D045726:Metalloproteases; D019158:N-Acetylneuraminic Acid