5KJX

Co-crystal Structure of PKA RI alpha CNB-B domain with cAMP

Summary for 5KJX

• Entry DOI: 10.2210/pdb5kjx/pdb
• Related: 5KJY 5KJZ
• Descriptor: cAMP-dependent protein kinase type I-alpha regulatory subunit, ADENOSINE-3',5'-CYCLIC-MONOPHOSPHATE (3 entities in total)
• Functional Keywords: cyclic nucleotide, camp-dependent protein kinase, nucleotide selectivity, cyclic nucleotide binding domain, signaling protein
• Biological source: Homo sapiens (Human)
• Total number of polymer chains: 1
• Total formula weight: 17285.71

Authors

Lorenz, R.,Moon, E.,Kim, J.J.,Huang, G.Y.,Kim, C.,Herberg, F.W. (deposition date: 2016-06-20, release date: 2017-06-28, Last modification date: 2023-09-27)

Primary citation

Lorenz, R.,Moon, E.W.,Kim, J.J.,Schmidt, S.H.,Sankaran, B.,Pavlidis, I.V.,Kim, C.,Herberg, F.W.
Mutations of PKA cyclic nucleotide-binding domains reveal novel aspects of cyclic nucleotide selectivity.
Biochem. J., 474:2389-2403, 2017

PubMed Abstract: Cyclic AMP and cyclic GMP are ubiquitous second messengers that regulate the activity of effector proteins in all forms of life. The main effector proteins, the 3',5'-cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA) and the 3',5'-cyclic guanosine monophosphate (cGMP)-dependent protein kinase (PKG), are preferentially activated by cAMP and cGMP, respectively. However, the molecular basis of this cyclic nucleotide selectivity is still not fully understood. Analysis of isolated cyclic nucleotide-binding (CNB) domains of PKA regulatory subunit type Iα (RIα) reveals that the C-terminal CNB-B has a higher cAMP affinity and selectivity than the N-terminal CNB-A. Here, we show that introducing cGMP-specific residues using site-directed mutagenesis reduces the selectivity of CNB-B, while the combination of two mutations (G316R/A336T) results in a cGMP-selective binding domain. Furthermore, introducing the corresponding mutations (T192R/A212T) into the PKA RIα CNB-A turns this domain into a highly cGMP-selective domain, underlining the importance of these contacts for achieving cGMP specificity. Binding data with the generic purine nucleotide 3',5'-cyclic inosine monophosphate (cIMP) reveal that introduced arginine residues interact with the position 6 oxygen of the nucleobase. Co-crystal structures of an isolated CNB-B G316R/A336T double mutant with either cAMP or cGMP reveal that the introduced threonine and arginine residues maintain their conserved contacts as seen in PKG I CNB-B. These results improve our understanding of cyclic nucleotide binding and the molecular basis of cyclic nucleotide specificity.

PubMed: 28583991
DOI: 10.1042/BCJ20160969

Experimental method

X-RAY DIFFRACTION (1.9 Å)