Role of a disulphide bond in Helicobacter pylori arginase.
Arginase is a binuclear Mn(2+)-metalloenzyme of urea cycle that hydrolyses arginine to ornithine and urea. Unlike other arginases, the Helicobacter pylori enzyme is selective for Co(2+). Previous study reported that DTT strongly inhibits the H. pylori enzyme activity suggesting that a disulphide bond is critical for the catalysis. In this study, we have undertaken steady-state kinetics, circular dichroism and mutational analysis to examine the role of a disulphide bond in this protein. By mutational analysis, we show that the disulphide bond is not important for catalytic activity; rather it plays an important role for the stability of the protein as observed from thermal denaturation studies. The loss of catalytic activity in the wild-type protein with DTT is due to the interaction with Co(2+). This is verified with the Mn(2+)-reconstituted proteins which showed a marginal loss in the activity with DTT.
Srivastava
Abhishek
A
National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110 067, India.
Dwivedi
Nidhi
N
Sau
Apurba Kumar
AK
D001119:Arginase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002384:Catalysis; D003035:Cobalt / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry* / Q000235:genetics; D004252:DNA Mutational Analysis; D004229:Dithiothreitol / Q000737:chemistry; D004795:Enzyme Stability; D016480:Helicobacter pylori / Q000201:enzymology*; D008345:Manganese / Q000378:metabolism; D009154:Mutation; D011489:Protein Denaturation / Q000235:genetics
D003035:Cobalt; D008345:Manganese; D001119:Arginase; D003545:Cysteine; D004229:Dithiothreitol