QUATERNARY STRUCTURES OF A CATALYTIC SUBUNIT-REGULATORY SUBUNIT DIMERIC COMPLEX AND THE HOLOENZYME OF THE CAMP-DEPENDENT PROTEIN-KINASE BY NEUTRON CONTRAST VARIATION

Chimeric molecules of the cAMP-dependent protein kinase (PKA) holoenzy me (R2C2) and of Delta(1-91)RC dimer were reconstituted using deuterat ed regulatory (R) and protiated catalytic (C) subunits. Small angle sc attering with contrast variation has revealed the shapes and dispositi ons of R and C in the reconstituted complexes, leading to low resoluti on models for both forms. The crystal structures of C and a truncation mutant of R fit well within the molecular boundaries of the RC dimer model. The area of interaction between R and C is small, seemingly poi sed for dissociation upon a conformational transition within R induced by cAMP binding. Within the RC dimer, C has a ''closed'' conformation similar to that seen for C with a bound pseudosubstrate peptide. The model for the PKA holoenzyme has an extended dumbbell shape. The inter connecting bar is formed from the dimerization domains of the R subuni ts, arranged in an antiparallel configuration, while each lobe contain s the cAMP-binding domains of one R interacting with one C. Our studie s suggest that the PKA structure may be flexible via a hinge movement of each dumbbell lobe with respect to the dimerization domain. Sequenc e comparisons suggest that this hinge might be a property of the R, PI CA isoforms.