Ala335 is essential for high‐affinity cAMP‐binding of both sites A and B of cAMP‐dependent protein kinase type I
- 10 April 1995
- journal article
- Published by Wiley in FEBS Letters
- Vol. 362 (3) , 291-294
- https://doi.org/10.1016/0014-5793(95)00261-7
Abstract
A single amino acid substitution (Ala335 Asp) in cAMP binding site B of the regulatory subunit of cAMP‐dependent protein kinase type I was sufficient to abolish high affinity cAMP binding for both cAMP binding sites A and B. Furthermore, the Ala335 Asp mutation increased the activation constant for cAMP of the mutant holoenzyme 30‐fold and also enhanced the rate of holoenzyme formation. Thus, the substitution was responsible for the dominant negative phenotype of the enzyme. Activation of mutant holoenzyme with site‐selective cAMP analogs indicated that the enzyme dissociated through binding to site A only. Our results provide evidence that Ala335 is an essential residue for high affinity cAMP binding of both sites as well as for the functional integrity of the enzyme.Keywords
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