INVESTIGADORES
ROSSI Silvia Graciela
congresos y reuniones científicas
Título:
STUDY OF THE INTERACTION BETWEEN THE R AND C SUBUNITS OF THE PROTEIN KINASE A IN THE FUNGUS MUCOR CIRCINELLOIDES
Autor/es:
OCAMPO J; MORENO S; ROSSI S.
Lugar:
Pinamar, Buenos Aires
Reunión:
Congreso; SAIB 41st Annual Meeting XLI Reunión Anual -Argentine Society for Biochemistry and Molecular Biology- -Sociedad Argentina de Investigación Bioquímica y Biología Molecular-; 2005
Institución organizadora:
SAIB-SAN
Resumen:
Protein kinase A is a tetramer composed of a dimer of regulatory
subunits (R) and two monomeric catalytic subunits (C). In the
holoenzyme C interacts with the pseudosubstrate site in R. This
pseudosubstrate site and additional contact sites in R render an
affinity in the nanomolar range. The affinities between R and C
subunits of PKA are quite different depending on the specie, finding
the highest values in M.rouxii and M.circinelloides.
Our hypothesis is that in the amino-termini region of R subunits
exist elements that interact with C subunits that are responsible for
the different affinity. The wild type R subunit and two mutant, R1
laking the N-terminal and R2 lacking a region of acidic residues
were cloned and expressed in a S.cerevisiae strain tpk2- tpk3- bcy1-
(133). Interaction between wtR or mutants R, and homologous or
heterologous C subunit was evaluated by in vivo and in vitro assays.
In vivo assays were made in yeast analyzing differences in
phenotypes and growth between the different transformants. In
vitro assays were made assaying the inhibition of C homologous or
heterologous by each R subunit and dissociation of holoenzymes
reconstituted. The IC50 for wtR was lower than those for mutant
Rs. Dissociation of reconstituted holoenzyme showed that the
concentration of cAMP required to dissociate the wtR-C
holoenzyme was higher than the concentration requeried to dissociate
the mutant R-C holoenzyme These results indicates that the acidic
residues region may be important in the high affinity interaction RC
found in M.rouxii and M.circinelloides PKA.