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MALDI mass sequencing and biochemical characterization of Setaria cervi protein tyrosine phosphatase
- Source :
- Parasitology Research. 112:147-154
- Publication Year :
- 2012
- Publisher :
- Springer Science and Business Media LLC, 2012.
-
Abstract
- A 30-kDa acid phosphatase with protein tyrosine phosphatase activity was identified in Setaria cervi (ScPTP). The enzyme was purified to homogeneity using three-step column chromatography. Matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) analysis of purified ScPTP yielded a total of eight peptides matching most closely to phosphoprotein phosphatase of Ricinus communis (RcPP). A hydrophilicity plot of RcPP revealed the presence of these peptides in the hydrophilic region, suggesting their antigenic nature. The substrate specificity of ScPTP with ortho-phospho-L-tyrosine and inhibition with sodium orthovanadate and ammonium molybdate affirmed it as a protein tyrosine phosphatase. ScPTP was also found to be tartrate resistant. The Km and Vmax were 6.60 mM and 83.3 μM/ml/min, respectively, with pNPP and 8.0 mM and 111 μM/ml/min, respectively, with ortho-phospho-L-tyrosine as the substrate. The Ki value with sodium orthovanadate was calculated to be 16.10 mM. Active site modification with DEPC, EDAC and pHMB suggested the presence of histidine, cysteine and aspartate at its active site. Thus, on the basis of MALDI-TOF and biochemical studies, it was confirmed that purified acid phosphatase is a PTP.
- Subjects :
- Protein tyrosine phosphatase
Biology
Substrate Specificity
chemistry.chemical_compound
Animals
Enzyme Inhibitors
Phosphotyrosine
Sodium orthovanadate
Filarioidea
Histidine
chemistry.chemical_classification
General Veterinary
Acid phosphatase
Active site
Substrate (chemistry)
General Medicine
Molecular biology
Kinetics
Infectious Diseases
Enzyme
Biochemistry
chemistry
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Insect Science
biology.protein
Parasitology
Protein Tyrosine Phosphatases
Cysteine
Subjects
Details
- ISSN :
- 14321955 and 09320113
- Volume :
- 112
- Database :
- OpenAIRE
- Journal :
- Parasitology Research
- Accession number :
- edsair.doi.dedup.....f8772a2785b01de111b57ac668519f61