?Role of the N ? terminal domain of the Regulatory Subunit of Protein Kinase A from fungi in dimerization and tethering to proteins?

MORENO, SILVIA; GALELLO, FIORELLA; RINALDI, JIMENA; GONZALEZ BARDECI, NICOLÁS; PEREYRA, ELBA; ROSSI, SILVIA

Oslo

Workshop; 3rd International Meeting on Anchored cAMP Signalling Pathways; 2010

Protein Society

Among fungi both ascomycetes and basidiomycetes have only one isoform of regulatory subunit (R) of PKA, while zygomycetes have four isoforms. A bioinformatic comparison of the divergent amino termini of the R subunits from fungi with those of mammals showed a conserved region with characteristics of dimerization domains (D/D) both in primary and secondary structure. A subfillum of ascomycetes, exemplified by Neurospora crassa, lacked this region and showed to have a monomeric R subunit. We studied in detail the N-terminus from Saccharomyces cerevisiae (ascomycete) and Mucor circinelloides (zygomycete). In the S.cerevisiae dimeric R (Bcy1) subunit the deletion of the D/D yielded a monomeric isoform.  A comparison of the D/D region of  Bcy1 and the four R subunits from M. circinelloides with the D/D domains of RI and RII predicts possible interacting surfaces for these subunits with other proteins. By pull-down assay Bcy1 was shown to interact with D-AKAP2. The addition of the permeable version of Ht-31 to M.circinelloides growth medium, had significant phenotypic consequences. Possible interactors of Bcy1 were selected making theoretical predictions of putative AKAPs from the yeast genome *.  Determinants and strength of the interactions with Bcy-1 and the role of the N-terminus in these interactions were assayed by peptide array. * In collaboration with W. Mac Laughlin from Susan Taylor?s lab