STUDY ON THE PROPERTIES OF PURIFIED RECOMBINANT SUPEROXIDE DISMUTASE FROM STAPHYLOCOCCUS EQUORUM, A LOCAL ISOLATE FROM INDONESIA
Journal Title: International Journal of Pharmacy and Pharmaceutical Sciences - Year 2014, Vol 6, Issue 11
Abstract
Objective: Superoxide dismutase (SOD) (E. C: 1.15.1.1) from Staphylococcus equorum which catalyzes the dismutation of the superoxide anion (O2.-) into molecular oxygen (O2) and hydrogen peroxide (H2O2), is one of the most important classes of antioxidant enzymes and are used in pharmaceutical or cosmetic applications. SOD of S. equorum was purified from total protein into homogeneity and characterized to determine the unit activity, ion metal cofactor, optimum temperature and pH, kinetic parameters, and effect of denaturing and reducing agents and UVC exposure on the rSOD activity.Methods: The protein was purified in a single-step purification using Ni-NTA afï¬nity column with various imidazole concentrations. SOD activity was analyzed by colorimetric and activity staining using nitroblue tetrazolium (NBT). The purified rSOD was exposed to different temperatures and pHs, different concentrations of denaturing agents, reducing agents, and to UVC exposure.Results: SOD protein with high purity was obtained when imidazole concentrations of 100 mM, 200 mM and 250 mM were applied. The purified rSOD displayed specific activity of 1666.7 U mg-1when measured at 30ºC and pH 7.8. The presence of conserved manganese-binding sites (H28, H83, D171, H175) and the inhibition of rSOD activity by NaN3 but not by H2O2 or KCN and indicated that rSOD was Mn-dependent. The optimum temperature and pH were determined to be 40ºC and 6.0, respectively. The Michaelis constant (Km), maximum velocity (Vmax), turnover number (kcat) and catalytic efficiency (kcat/Km) were found to be 371.2 µM, 1.738 µMS-1, 1.358 s-1, and 3.7x10-3 S-1µM-1, respectively. The rSOD activity was slightly affected in the presence of detergents (0.5% SDS, 0.5% Triton-X 100), denaturing agents (6 M GdnHCl and 6 M urea) and reducing agent (5 mM βME). After exposure of rSOD by UVC for 45 min, it retained half of its activity.Conclusion: This is the first study to report the stability of the SOD of S. equorum against environmental factors. The SOD displays some thermostability, is active in wide pH, stable in the presence of denaturing and reducing agents, however it is relatively unstable to UVC exposure
Authors and Affiliations
Ana Indrayati, Sukmadjaja Asyarie, Tri Suciati, Debbie Soefie Retnoningrum
Keywords
Related Articles
- EP ID EP579337
- DOI -
- Views 52
- Downloads 0
BIOCHEMICAL CHARACTERIZATION AND ANTIBACTERIAL PROPERTIES OF FISH SKIN MUCUS OF FRESH WATER FISH, HYPOPHTHALMICHTHYS NOBILIS
Objective: The present study was undertaken to characterize the biochemical composition and antibacterial activity of skin mucus of fish Hypophthalmichthys nobilis against different human and fish pathogenic bacterial st...
MITIGATING ROLE OF ZINC AND IRON AGAINST CADMIUM INDUCED TOXICITY IN LIVER AND KIDNEY OF MALE ALBINO RAT: A STUDY WITH REFERENCE TO METALLOTHIONEIN QUANTIFICATION
Objective: The present study is carried out to know the mitigating role of zinc (Zn) and / or iron (Fe) supplementation on cadmium (Cd) induced toxicity in rats with special reference to metallothionein (MT) protein.Meth...
ECO-FRIENDLY NATURAL ACID-BASE INDICATOR PROPERTIES OF FOUR FLOWERING PLANTS FROM WESTERN GHATS
Objective: The present study aims to evaluate the indicator property of ethanolic extract of the flowers of the Quisqualis indica L, Pentas lanceolata, Melastoma malabathricum L and Impatiens acaulis Methods: Four differ...
VALIDATION OF ACCUMULATION OF CAMPTOTHECIN CONTENT, AN ANTI-CANCER ALKALOID IN NOTHAPODYTES NIMMONIANA GRAHAM. IN PHENOTYPIC VARIANTS: METHOD FOR IDENTIFYING HIGH-YIELDING SOURCES OF ALKALOID
Objective: The study was aimed to find the CPT accumulation, in particular organ, via variable extraction method and in vitro propagation method both qualitatively and quantitatively.Methods: In the present study, the se...
SIMULTANEOUS QUANTIFICATION OF BUPRENORPHINE HCL AND NALOXONE HCL BY VIERORDT’S METHOD
Objective: Development of a simple, rapid and accurate vierordt’s method or simultaneous equation (SE) method for the simultaneous estimation of Buprenorphine HCl (BU) and Naloxone HCl (NA) in bulk and pharmaceutical d...