Article
ADPase activity of recombinantly expressed thermotolerant ATPases may be caused by copurification of adenylate kinase of Escherichia coli
The FEBS Journal
(2009)
Abstract
Except for apyrases, ATPases generally target only the γ‐phosphate of a nucleotide. Some non‐apyrase ATPases from thermophilic microorganisms are reported to hydrolyze ADP as well as ATP, which has been described as a novel property of the ATPases from extreme thermophiles. Here, we describe an apparent ADP hydrolysis by highly purified preparations of the AAA+ ATPase NtrC1 from an extremely thermophilic bacterium, Aquifex aeolicus. This activity is actually a combination of the activities of the ATPase and contaminating adenylate kinase (AK) from Escherichia coli, which is present at 1/10 000 of the level of the ATPase. AK catalyzes conversion of two molecules of ADP into AMP and ATP, the latter being a substrate for the ATPase. We raise concern that the observed thermotolerance of E. coli AK and its copurification with thermostable proteins by commonly used methods may confound studies of enzymes that specifically catalyze hydrolysis of nucleoside diphosphates or triphosphates. For example, contamination with E. coli AK may be responsible for reported ADPase activities of the ATPase chaperonins from Pyrococcus furiosus, Pyrococcus horikoshii, Methanococcus jannaschii and Thermoplasma acidophilum; the ATP/ADP‐dependent DNA ligases from Aeropyrum pernix K1 and Staphylothermus marinus; or the reported ATP‐dependent activities of ADP‐dependent phosphofructokinase of P. furiosus. Purification methods developed to separate NtrC1 ATPase from AK also revealed two distinct forms of the ATPase. One is tightly bound to ADP or GDP and able to bind to Q but not S ion exchange matrixes. The other is nucleotide‐free and binds to both Q and S ion exchange matrixes.
Keywords
- AAA+ ATPase,
- adenylate kinase,
- ADPase; σ54,
- thermophilic proteins
Disciplines
Publication Date
February, 2009
DOI
10.1111/j.1742-4658.2008.06825.x
Publisher Statement
This is the accepted version of the following article: Chen, Baoyu, Tatyana A. Sysoeva, Saikat Chowdhury, Liang Guo, and B. Tracy Nixon. "ADPase activity of recombinantly expressed thermotolerant ATPases may be caused by copurification of adenylate kinase of Escherichia coli." The FEBS Journal 276, no. 3 (2009): 807-815, which has been published in final form at doi: 10.1111/j.1742-4658.2008.06825.x.
Citation Information
Baoyu Chen, Tatyana A. Sysoeva, Saikat Chowdhury, Liang Guo, et al.. "ADPase activity of recombinantly expressed thermotolerant ATPases may be caused by copurification of adenylate kinase of Escherichia coli" The FEBS Journal Vol. 276 Iss. 3 (2009) p. 807 - 815 Available at: http://works.bepress.com/baoyu-chen/8/