Partial purification and kinetic properties of Catalase from leaves of Withania somnifera compared under copper stress
Dr. Anupama Singh
, Prof.(Dr.) Kapil Lawrence
Catalase, DEAE-cellulose chromatography, Hydrogen peroxide, Sodium azide, SDS-PAGE, kinetic constants and Withania somnifera.
The leaves of control (0 mM) and Cu2+ treated (50 mM) plant were used to study partial purification, characterization and kinetic properties of the catalase. The native polyacrylamide gel electrophoresis (PAGE) emphasize that band of catalase (EC 1.11.1.6) were expressed with maximum intensity in Cu2+ treated (50 mM) Withania somnifera leaves. The partial purification includes fractionation by 0-40% ammonium sulphate [(NH4)2SO4] and elution by DEAE-cellulose chromatography. The enzyme was approximately 14-fold purified with a final overall yield of 73.93 % to a specific activity of 7.57 Umg-1 proteins in the control leaves extract. However, from Cu2+ treated (50 mM) leaves extracts catalase were partially purified 30-fold with overall yield 72.67 % to a specific activity of 19.67 Umg-1 proteins. Through SDS-PAGE (10%) the mol. wt. of the catalase subunit obtained as ≈60 KDa, possibly may be tetramer. catalase activity was optimum at pH 7.0 in 0.1M phosphate buffer at a temperature ranged from 35°C-45°C. From Lineweaver-Burk plot, enzyme with substrate hydrogen peroxide (H2O2) yielded an apparent Km and Vmax of 31.29 mM and 585 µM min-1 from control leaves; however, 56.60 mM and 1223 µM min-1 from 50 mM Cu2+ treated leaves respectively. The kinetic constant, Ki obtained from Dixon plots with inhibitor NaN3 were 1.3 mM and 1.7 mM for control and Cu2+ treated (50 mM) respectively. The Dixon plots revealed a competitive type of inhibition for catalase with substrate H2O2 against sodium azide (NaN3) as inhibitor.
"Partial purification and kinetic properties of Catalase from leaves of Withania somnifera compared under copper stress", IJSDR - International Journal of Scientific Development and Research (www.IJSDR.org), ISSN:2455-2631, Vol.9, Issue 6, page no.310 - 326, June-2024, Available :https://ijsdr.org/papers/IJSDR2406036.pdf
Volume 9
Issue 6,
June-2024
Pages : 310 - 326
Paper Reg. ID: IJSDR_211641
Published Paper Id: IJSDR2406036
Downloads: 000347094
Research Area: Life Sciences
Country: Varanasi, Uttar-Pradesh, India
ISSN: 2455-2631 | IMPACT FACTOR: 9.15 Calculated By Google Scholar | ESTD YEAR: 2016
An International Scholarly Open Access Journal, Peer-Reviewed, Refereed Journal Impact Factor 9.15 Calculate by Google Scholar and Semantic Scholar | AI-Powered Research Tool, Multidisciplinary, Monthly, Multilanguage Journal Indexing in All Major Database & Metadata, Citation Generator
Publisher: IJSDR(IJ Publication) Janvi Wave
