Key result
The R14 deletion in phospholamban reduces its binding affinity to protein kinase A and dramatically reduces phosphorylation kinetics by preventing the enzyme from adopting catalytically committed conformations.
The R14 deletion in phospholamban impairs its phosphorylation by protein kinase A by altering the enzyme's conformational dynamics, providing a molecular mechanism for its role in familial dilated cardiomyopathy.
NMR dynamics differ in R14-deleted phospholamban-PKA complexes; leaves open in vivo catalytic and cardiac effects.
The dynamic interplay between kinases and substrates is crucial for the formation of catalytically committed complexes that enable phosphoryl transfer. However, a clear understanding on how substrates modulate kinase structural dynamics to control catalytic efficiency is still missing. Here, we used solution NMR spectroscopy to study the conformational dynamics of two complexes of the catalytic subunit of the cAMP-dependent protein kinase A with WT and R14 deletion phospholamban, a lethal human mutant linked to familial dilated cardiomyopathy. Phospholamban is a central regulator of heart muscle contractility, and its phosphorylation by protein kinase A constitutes a primary response to β-adrenergic stimulation. We found that the single deletion of arginine in phospholamban's recognition sequence for the kinase reduces its binding affinity and dramatically reduces phosphorylation kinetics. Structurally, the mutant prevents the enzyme from adopting conformations and motions committed for catalysis, with concomitant reduction in catalytic efficiency. Overall, these results underscore the importance of a well-tuned structural and dynamic interplay between the kinase and its substrates to achieve physiological phosphorylation levels for proper Ca(2+) signaling and normal cardiac function.
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Kim et al. (2015) studied Dilated cardiomyopathy (molecular mechanism). PLN R14del mutant vs. PLN WT was evaluated on Phosphorylation kinetics and binding affinity. The R14 deletion in phospholamban reduces its binding affinity to protein kinase A and dramatically reduces phosphorylation kinetics by preventing the enzyme from adopting catalytically committed conformations.
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