Purification and Characterisation of isozymes 1 and 2 of Acid phosphatase from Cicer arietinum
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Purification & Characterisation of isozyme-1 and 2 of Acid Phosphatase from Cicer arietinum Antik K Bose Author information Affilations Human Biology Division, Fred Hutchinson Cancer Research Center, Seattle, USA. Antik K Bose Biochemical properties of acid phosphatase isoforms obtained from germinating seeds of Cicer arietinum have been described. The isoforms were purified 3604.88 and 2390 fold with 1.4% recovery by CM-Cellulose, DEAE- Cellulose, (L)-tartarate-AH-Sepharose-4B affinity chromatography and Isoelectric focusing. The molar mass of both isoforms was estimated by Sephadex G-200 to be 100KD and by 6% SDS- PAGE a band of 55KD was obtained suggesting a homodimer. Temperature optima for both isoforms is 32 C and pH optima are 4.5 and 5.5 for isoforms 1 and 2. The isoforms were moderately glycosylated , 40% and 35% saccharide content for isoforms 1 and 2. Isoform-1 dephosphorylated Phosphotyrosine, PNPP, o-phospho-L- serine, Phosphoenolpyruvate, PPi and ATP. Both isoforms showed product inhibition by phosphate. The activity was enhanced by Mg2 + ( 2.2% for isoform-1 and 4.9% for isoform-2 ). 4-nitrophenyl phosphate was hydrolysed with K M and V max 2,27 mM and 238.095 μM PNPP consumed/min for isoform-2. Vanadium tetrachloride showed uncompetitive inhibition on isoform-1 and competitive inhibition on isoform-2. Nature Methods ISSN: 1548-7091 EISSN: 1548-7105 Journal home | Current issue | Archive | Press releases | ©2008 Nature Publishing Group | Privacy policy
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Transcript of Purification and Characterisation of isozymes 1 and 2 of Acid phosphatase from Cicer arietinum
Purification & Characterisation of isozyme-1 and 2 of Acid Phosphatase from Cicer arietinum
Antik K Bose
Author information
Affilations
Human Biology Division, Fred Hutchinson Cancer Research Center, Seattle, USA.
Antik K Bose
Biochemical properties of acid phosphatase isoforms obtained from germinating seeds of Cicer arietinum have been described. The
isoforms were purified 3604.88 and 2390 fold with 1.4% recovery by CM-Cellulose, DEAE- Cellulose, (L)-tartarate-AH-Sepharose-4B affinity
chromatography and Isoelectric focusing. The molar mass of both isoforms was estimated by Sephadex G-200 to be 100KD and by 6% SDS-
PAGE a band of 55KD was obtained suggesting a homodimer. Temperature optima for both isoforms is 32 C and pH optima are 4.5 and 5.5
for isoforms 1 and 2. The isoforms were moderately glycosylated , 40% and 35% saccharide content for isoforms 1 and 2. Isoform-1
dephosphorylated Phosphotyrosine, PNPP, o-phospho-L- serine, Phosphoenolpyruvate, PPi and ATP. Both isoforms showed product
inhibition by phosphate. The activity was enhanced by Mg2+ ( 2.2% for isoform-1 and 4.9% for isoform-2 ). 4-nitrophenyl phosphate was
hydrolysed with KM and Vmax 2,27 mM and 238.095 µM PNPP consumed/min for isoform-2. Vanadium tetrachloride showed uncompetitive
inhibition on isoform-1 and competitive inhibition on isoform-2.
Nature Methods
ISSN: 1548-7091
EISSN: 1548-7105
Journal home | Current issue | Archive | Press releases |
©2008 Nature Publishing Group | Privacy policy
