Simple and fast purification of Escherichia coli adenylate kinase

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Abstract
Adenylate kinase from E. coli (strains CR341 and CR341 T28, a temperature‐sensitive mutant) was purified by a two‐step chromatographic procedure. The enzyme from crude extracts of both mutant and parent strain was bound to blue—Sepharose at pH 7.5, thereafter specifically eluted with 0.05 mM P 1,P 5‐di(adenosine‐5′)pentaphosphate. A second chromatography on Sephadex G‐100 yielded pure enzyme. E. coli adenylate kinase was strongly inhibited by P 1,P 5‐di(adenosine‐5′)pentaphosphate (K i 0.6 μM for adenylate kinase of strain CR341 and 2.1 μM in the case of mutant enzyme). After denaturation in 6 M guanidinium hydrochloride both mutant and parent adenylate kinase returned rapidly to the native, active state by dilution of guanidinium hydrochloride.