Identification and characterization of novel membrane-bound PRL protein tyrosine phosphatases from Setaria cervi, a bovine filarial parasite.
A significant amount of protein tyrosine phosphatase (PTP) activity was detected in the detergent-soluble membrane-bound fraction of Setaria cervi, a bovine filarial parasite. The membrane-bound PTP activity was significantly inhibited when the adult parasites were exposed to compounds having antifilarial activity like aspirin and SK7 as well as phenylarsine oxide, a specific PTP inhibitor suggesting that this activity is stress regulated. Further, this enzyme was purified as a single protein of apparently 21 kDa using two different chromatographic techniques. The MALDI-MS/MS analysis of its peptides showed closest match with protein tyrosine phosphatase PRL (Aedes aegypti). This purified enzyme (named as PRL) showed maximum activity at pH 5.5/37 °C and hydrolysed para nitro phenyl phosphate (pNPP) at the highest rate followed by O-P-L-tyrosine and O-P-L-threonine. It showed significant inhibition by specific inhibitors of PTP such as sodium orthovanadate, phenylarsine oxide and ammonium molybdate and was activated by dithiothreitol (DTT). The active site modification studies suggested involvement of cysteine, arginine, histidine and aspartic acid in the catalytic activity of PRL. The activity of S. cervi PRL was also found to be resistant towards the external oxidative stress. Thus, S. cervi PRL could be taken as a potential target for the management of human lymphatic filariasis.
Department of Biochemistry, Faculty of Science, Banaras Hindu University, Varanasi, 221005, India.
Neetu
N
Singh
Department of Biochemistry, Faculty of Science, Banaras Hindu University, Varanasi, 221005, India.
Smita
S
Yadav
Department of Biochemistry, Faculty of Science, Banaras Hindu University, Varanasi, 221005, India. surathaur@rediffmail.com.
Sushma
S
Rathaur
D000818:Animals; D020134:Catalytic Domain; D002417:Cattle; D002418:Cattle Diseases / Q000469:parasitology*; D015801:Helminth Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D017027:Protein Tyrosine Phosphatases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017182:Setaria Nematode / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D012719:Setariasis / Q000469:parasitology*; D053719:Tandem Mass Spectrometry
D015801:Helminth Proteins; D017027:Protein Tyrosine Phosphatases
MALDI-MS/MS; Protein tyrosine phosphatase; Setaria cervi