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Purification and Characterization of a Thermostable Pyruvate Ferredoxin Oxidoreductase/Pyruvate Decarboxylase from Thermococcus kodakaraensis

Purification and Characterization of a Thermostable Pyruvate Ferredoxin Oxidoreductase/Pyruvate Decarboxylase from Thermococcus kodakaraensis

Kanwal Nisa1, Sadaf Ashraf1, Masood Ahmed Siddiqui1*,Naila Taj1Habib-Ur-Rehman1, Arifa Bano1 and Naeem Rashid2

1Department of Chemistry, Biotechnology Research Laboratory, University of Balochistan, Quetta, 87300, Pakistan 
2School of Biological Sciences, University of the Punjab, Quaid-e-Azam Campus, Lahore 54590, Pakistan

*      Corresponding author: masoodbioeng@hotmail.com

 

ABSTRACT

Thermococcus kodakaraensis is a strictly anaerobic sulfur dependent archaeon that grows optimally at 85oC by a fermentative type metabolism. An extremely thermostable bifunctional enzyme, which exhibits pyruvate ferredoxin oxidoreductase (POR) and pyruvate decarboxylase (PDC) activities, was purified to an apparent homogeneity from this archaeon. The purified enzyme exhibited optimal activities at 95oC for POR and 90oC for PDC reactions. The optimum pH for POR reaction was 8.5 and for that of PDC it was 9.5, in the absence of oxygen. The specific activities for POR and PDC reactions were 22 U/mg and 3.8 U/mg, respectively. The native enzyme had an apparent molecular weight of 240 kDa and was a dimer of heterotetramers (αβɣδ)2 with molecular masses of 44, 36, 20 and 12 kDa, respectively. Both of the activities were oxygen sensitive. The apparent Km values for POR reaction towards pyruvate and CoASH were 0.49 µM and 115 µM, respectively, while for PDC reaction values these values were µM 0.34 and 42 µM. To the best of our knowledge this is the first report on purification and characterization of a POR/PDC from T. kodakaraensis.
 

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Pakistan Journal of Zoology

June

Vol. 52, Iss. 3, Pages 825-1224

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