Regulatory Role Of Arginine Deiminase In The Biology Of The Primitive Eukaryote Giardia lamblia.

TOUZ MC; RIVERO MR; ROPOLO AS; CONRAD JT; SVARD S; NASH TE.

Morelia, Michoacán Mexico.

Congreso; II International Giardia and Cryptosporidium Conference.; 2007

Giardia lamblia undergoes antigenic variation, a process where a single variant-specific surface protein (VSP) is replaced by an antigenically-different VSP. All VSP genes (~150) encode type I integral membrane proteins that have a conserved five-amino acid cytoplasmic tail, CRGKA. To identify molecules interacting with this tail, the peptide His6-CRGKA was employed in pull-down assays. Surprisingly, arginine deiminase (ADI) bound specifically to CRGKA as identified by mass spectroscopy.  In Giardia and prokaryotes, ADI is known to catalyze the irreversible catabolism of free arginine to citrulline in the arginine dihydrolase pathway as an important source of energy. In contrast, higher eukaryotes possess peptidil-arginine deiminases (PAD), enzymes that post-translationally deiminates the guanidine group of arginine residues in peptides and proteins to generate ammonia and citrulline. This post-translational modification has a large impact in the structure and function of the target protein. Analysis of Giardia proteins using a detection method that is specific for the presence of citrulline, showed that VSPs are citrullinated. Activity analysis using His6-CRGKA as substrate and purified gADI, demonstrated that this peptide can be citrullinated in vitro (CcitGKA). Immunofluorescence analysis of gADI showed cytoplasmic localization and close association to the plasma membrane. When trophozoites were induced to differentiate into cysts, gADI translocated from the cytoplasm to the nuclei suggesting that ADI has a specific role in the process of encystation. These results show that citrullination may play an essential role in both defense mechanisms of Giardia, encystation and antigenic variation.