Question

Research in cell biology and metabolism has progressed due to the discovery of molecules that artificially stimulate or inhibit glucagon/epinephrine and insulin signaling pathways. Let’s say you are working in a lab cataloging the effects of a library of small molecules on these pathways and have a “hit” on molecule 1stAVNGR. Preliminary data on molecule 1stAVNGR indicates that the cardiac isoform of PFK2/FBPase2 is doubly phosphorylated when this molecule is present at micromolar concentrations in cell cultures. Given this context answer the following questions.

Under these conditions what is the predicted degree of association between the regulatory subunits and the catalytic subunits of PKA?

Answer 1

ANS.The Protein kinase A is clasified as type I or II depends on the association of catalytic subunit with Regulatory subunits.the cAMP- dependent protein kinase from Candida albicans is tetramer composed of two catalytic subunits(C) and two type II regulatory subunits(R).to evaluate the role of a putative autophosphorylation site of the R subunit in the interaction with C,the site was mutated to an Ala residue.Recombinant wild type and mutant forms of the R-subunit were expressed in E.coli.and purified.the wild type recombinant R subunit was fully phosphorylated by the purified C subunit .disassociation of mutant holoenzyme by cAMP was not affectd by the Mg ATP.metabolic labelling of yeast cells with orthophosphate indicated that the R subunit exists as a serine phophorylated protein.