Structural, enzymatic and biochemical studies on Helicobacter pylori arginase

Abstract

Arginase is an enzyme involved in the last step of the urea cycle, where it catalyses the hydrolysis of l-arginine to generate l-ornithine and urea. Compared to the well-characterised arginases from animals, yeast and other bacteria, Helicobacter pylori arginase, or RocF, is unique in at least three aspects. Firstly, it has been identified as an important factor in evasion of the host's immune system and thus contributes to persistent infection by the bacterium. Secondly, the optimal catalytic conditions of RocF are different from those of other arginases. Finally, sequence alignment indicates that RocF possesses considerable differences at its N- and C-terminal from other arginases and harbours an insertion of 13 residues in the middle of the sequence. To better understand these unique biochemical and enzymatic properties, we therefore have embarked on determining the structure of RocF. In this study, the crystal structure of RocF was solved with the molecular replacement method. Based on the structure and systematic mutagenesis studies, we confirmed that the inserted residues form a helix that was not observed in other arginases and was able to raise the arginase activity by 30% probably by change the conformation of the substrate binding pocket. Six residues were involved in Mn2+ binding, all of which were essential for arginase activity. The C-terminal motif is not sufficient in establishing the oligomeric state of RocF, and no disulphide bonds were observed in RocF.