Identification and characterization of oxidoreductase component (NdmD) of methylxanthine oxygenase system in Pseudomonas sp. NCIM 5235

Applied Microbiology and Biotechnology
Sreeahila Retnadhas, Sathyanarayana N Gummadi

Abstract

Pseudomonas sp. NCIM 5235 is a caffeine-degrading bacterial strain that metabolizes caffeine by sequential demethylation using methylxanthine demethylases. These enzymes belong to the class of two-component Rieske oxygenases and require an oxidoreductase, NdmD, for efficient catalysis. NdmD in Pseudomonas sp. has a unique domain fusion in its N-terminal that is not observed in any other Rieske oxygenase reductases reported so far. In this report, a ~ 1.7 kb ndmD gene from the gDNA of Pseudomonas sp. has been isolated and has been cloned in a pET28a expression vector. Soluble NdmD was over-expressed in Escherichia coli BL21 cells and purified by Ni2+ NTA chromatography. Monomeric molecular mass of the protein was found to be ~ 65 kDa and optimal activity was observed at 35 °C and pH 8.0. It showed broad substrate specificity with highest Kcat/km of 490.8 ± 17.7 towards cytochrome c. To determine the role of N-terminal Rieske domain in its reductase activity, two deletion constructs Δ114NdmD and Δ250NdmD were made. Cytochrome c reductase (ccr) activity of the NdmD constructs and demethylase activity of NdmA in the presence of NdmD constructs showed that there is no significant difference in the catalytic activity of NdmD upon del...Continue Reading

References

Nov 1, 1995·FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology·G T GassnerD P Ballou
May 8, 2001·Archives of Biochemistry and Biophysics·C C Marohnic, M J Barber
Jun 8, 2002·Journal of Molecular Biology·Andreas KarlssonS Ramaswamy
Sep 26, 2006·International Journal of Food Microbiology·S GokulakrishnanSathyanarayana N Gummadi
Sep 30, 2006·Biotechnology Letters·Swati Sucharita Dash, Sathyanarayana N Gummadi
Sep 4, 2007·Proceedings of the National Academy of Sciences of the United States of America·Jinzhong LinJinfeng Wang
Aug 23, 2008·Journal of Basic Microbiology·Swati Sucharita Dash, Sathyanarayana N Gummadi
Dec 15, 2010·Journal of Bacteriology·Jeffery M BoydJohn W Peters
May 17, 2011·Applied Microbiology and Biotechnology·Sathyanarayana N Gummadi, B Bhavya
Feb 14, 2012·Journal of Bacteriology·Ryan M SummersMani Subramanian
Jun 5, 2012·The Journal of Biological Chemistry·Thanawat PhongsakPimchai Chaiyen
Feb 14, 2015·Microbial Biotechnology·Ryan M SummersMani Subramanian
Jul 15, 2015·Nature Communications·Javier A Ceja-NavarroEoin L Brodie

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Methods Mentioned

BETA
reductase assay
gel filtration

Software Mentioned

Origin
iTASSER
GraphPad Prism
TASSER
ExPASy PROSITE
SWISS MODEL

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