132
Modeling of Biomolecular Systems Interactions, Dynamics, and Allostery Poster Session II
79-POS
Board 32
Structural Features of the Gamma Subunit (AGG2) and Specificity of A. Thaliana
Heterotrimeric G-protein Interactions
Zehra Sayers
, Sandra Quarantini, Hazal B. Kose, Burak Gur.
Sabanci University, Istanbul, Turkey.
Heterotrimeric G-proteins are involved in several essential signaling pathways in plants,
including those controlling seed germination, growth and defense against pathogens. Despite
detailed functional and structural information on mammalian G-proteins, structure-function
characterization of the plant proteins is lacking. Moreover, although there are basic similarities in
the signaling mechanisms, regulation of activity in the mammalian and plant G-proteins show
important differences. We cloned and expressed A. thaliana subunits alpha, beta and gamma in
E. coli for structural studies and here results on the gamma subunit AGG2 will be presented.
Purified recombinant AGG2 shows a propensity to oligomerize in solution and as determined by
SDS- and native-PAGE, size exclusion chromatography and dynamic light scattering a stable
dimer is obtained only under fully reducing conditions. Small angle X-ray solution scattering
(SAXS) measurements reveal that AGG2 dimer has an elongated flexible structure which can be
modeled with the ensemble optimization method (EOM) used for intrinsically disordered
proteins. Results indicate that AGG2 structure can be described as a population of inter-
converting flexible conformers which all contain a rigid α-helix domain involved in the inter-
molecular coiled coil interactions. Estimation of the secondary structure content from circular
dichroism (CD) measurements agrees well with that predicted by homology modeling and SAXS
models. Results on thermal stability of the protein monitored by CD measurements are also in
agreement with the flexible structure predicted by EOM models. Structural models together with
sequence analyses show the centrality of the AGG2 N-terminal region in establishing the coiled-
coil interaction, crucial for binding to its natural partner AGB1. Our results also suggest that the
disordered flexible regions may be part of the mechanism for conferring functional selectivity to
the A. thaliana beta-gamma dimer at different location in the plant.
