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Structural and catalytic role of arginine 88 in Escherichia coli adenylate kinase as evidenced by chemical modification and site-directed mutagenesis

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Reinstein,  Jochen
Department of Biomolecular Mechanisms, Max Planck Institute for Medical Research, Max Planck Society;
Molecular chaperones, Max Planck Institute for Medical Research, Max Planck Society;

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Wittinghofer,  Alfred
Emeritus Group Biophysics, Max Planck Institute for Medical Research, Max Planck Society;

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Citation

Reinstein, J., Gilles, A., Rose, T., Wittinghofer, A., Saint−Girons, I., Barzu, O., et al. (1989). Structural and catalytic role of arginine 88 in Escherichia coli adenylate kinase as evidenced by chemical modification and site-directed mutagenesis. The Journal of Biological Chemistry, 264(14), 8107-8112. Retrieved from https://www.ncbi.nlm.nih.gov/labs/articles/2542263/.


Cite as: http://hdl.handle.net/11858/00-001M-0000-0019-ADA9-C
Abstract
Phenylglyoxal inactivates Escherichia coli adenylate kinase by modifying a single arginine residue (Arg-88). ATP, ADP, P1,P5-di(adenosine 5')-pentaphosphate, and to a lesser extent AMP protect the enzyme against inactivation by phenylglyoxal. Site-directed mutagenesis of Arg-88 to glycine yields a modified form of adenylate kinase (RG88 mutant) closely related structurally to the wild-type protein as indicated by Fourier transform infrared spectroscopy, differential scanning calorimetry, and limited proteolysis. However, this modified protein has only 1% of the maximum catalytic activity of the wild-type enzyme and 5- and 85-fold higher apparent Km values for ATP and AMP, respectively, than the parent adenylate kinase. Arg-88, which is a highly conserved residue in all known molecular forms of adenylate kinases (corresponding to Arg-97 in muscle cytosolic enzyme), should be located inside a big cleft of the molecule, close to the phosphate-binding loop. It possibly stabilizes the transferable gamma-phosphate group from ATP to AMP in the transition state.