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Isolation and Characterization of a Molybdenum Reducing Enzyme in Enterobacter cloacae Strain 48


Citation

Abd. Shukor, Mohd. Yunus and Lee, C. H. and Omar, Ismail and Karim, M. I. A. and Syed, Mohd Arif and Shamaan, Nor Aripin (2003) Isolation and Characterization of a Molybdenum Reducing Enzyme in Enterobacter cloacae Strain 48. Pertanika Journal of Science & Technology, 11 (2). pp. 261-272. ISSN 0128-7680

Abstract / Synopsis

Molybdenum reducing enzyme was isolated from Enterobacter cloacae Strain 48 by ammonium sulphate fractionation, DE-cellulose ion-exchange chromatography and Sephacryl S-200 gel filtration. SDS-PAGE of the concentrated Sephacryl S-200 gel filtration eluates revealed the presence of 3 protein subunits of molecular weight 80, 90 and 100 kDa. The active concentrated fraction from the Sephacryl S-200 gel filtration step was then characterized for molybdenum reducing activity with 12-molybdophosphate (12-MoP) as a substrate. The optimum pH and temperature of the reaction was 5.0 and 28-33°C, respectively. ADH was a better reducing agent in the reaction than NADPH; the double reciprocal plot of activity against ADH and NADPH revealed apparent Km and V""", values of 1.65 mM, 6.28 nmole molybdenum blue produced/min/mg and 2.13 mM and 4.10 nmole molybdenum blue produced/min/mg, respectively. The double reciprocal plot of activity against 12-MoP and 20-molybdodiphosphate revealed apparent K m values of 0.3 mM and 0.4 mM, respectively. The apparent Vmax values are similar for both substrates at 6 nmole molybdenum blue produced/min. The assay method for molybdenum reducing activity using 12-MoP was found to be easier and more rapid than the present method of using molybdate as a substrate


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Additional Metadata

Item Type: Article
Publisher: Universiti Putra Malaysia Press
Keywords: Molybdenum reducing enzyme, 12-m.olybdophosphate
Depositing User: Nur Izyan Mohd Zaki
Date Deposited: 01 Dec 2009 07:16
Last Modified: 27 May 2013 07:11
URI: http://psasir.upm.edu.my/id/eprint/3778
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