Purification and characterization of a novel caffeine oxidase from Alcaligenes species

Journal of Biotechnology
B R MohapatraA Mazumder

Abstract

Alcaligenes species CF8 isolated from surface water of a lake produced a novel serine type metallo-caffeine oxidase. The optimal medium for caffeine oxidase production by this strain was (w/v) NaNO(3), 0.4%; KH(2)PO(4), 0.15%; Na(2)HPO(4), 0.05%; FeCl(3).6H(2)O, 0.0005%; CaCl(2).2H(2)O, 0.001%; MgSO(4).7H(2)O, 0.02%; glucose, 0.2%; caffeine, 0.05%, pH 7.5. The enzyme was purified to 63-fold by using ammonium sulfate precipitation, dialysis, ion exchange (diethylaminoethyl-cellulose) and gel filtration (Sephadex G-100) chromatographic techniques. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that the purified caffeine oxidase was monomeric with a molecular mass of 65 kDa. The purified caffeine oxidase with a half-life of 20 min at 50 degrees C had maximal activity at pH 7.5 and 35 degrees C. The purified caffeine oxidase had strict substrate specificity towards caffeine (K(m) 8.94 microM and V(max) 47.62 U mg protein(-1)) and was not able to oxidize xanthine and hypoxanthine. The enzyme activity was not inhibited by para-chloromercuribenzoic acid, iodoacetamide, n-methylmaleimide, salicylic acid and sodium arsenite indicating the enzyme did not belong to xanthine oxidase family. The enzyme was not affected b...Continue Reading

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Citations

May 17, 2011·Applied Microbiology and Biotechnology·Sathyanarayana N Gummadi, B Bhavya
Dec 6, 2011·Applied Microbiology and Biotechnology·Sathyanarayana N GummadiNandhini Ashok
Feb 21, 2009·Journal of Industrial Microbiology & Biotechnology·Sathyanarayana N GummadiSanthosh Devarai
May 25, 2010·Journal of Industrial Microbiology & Biotechnology·Sathyanarayana N Gummadi, Devarai Santhosh
Sep 30, 2006·Biotechnology Letters·Swati Sucharita Dash, Sathyanarayana N Gummadi
Feb 14, 2015·Microbial Biotechnology·Ryan M SummersMani Subramanian
Apr 14, 2016·Environmental Science and Pollution Research International·Dongqing ZhangWun Jern Ng
Mar 12, 2008·Applied Microbiology and Biotechnology·Chun Ming WongXiao Dong Chen
Aug 23, 2008·Journal of Basic Microbiology·Swati Sucharita Dash, Sathyanarayana N Gummadi
Feb 6, 2020·Applied Microbiology and Biotechnology·Di SunZiqing Gao
Mar 11, 2020·Environmental Technology·Khirbet López-VelázquezDavid Herrera-López

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