Interconnecting Flexibility, Structural Communication, and Function in RhoGEF Oncoproteins

Dbl
family Rho guanine nucleotide exchange factors (RhoGEFs) play
a central role in cell biology by catalyzing the exchange of guanosine
5′-triphosphate for guanosine 5′-diphosphate (GDP) on
RhoA. Insights into the oncogenic constitutive activity of the Lbc
RhoGEF were gained by analyzing the structure and dynamics of the
protein in different functional states and in comparison with a close
homologue, leukemia-associated RhoGEF. Higher intrinsic flexibility,
less dense and extended structure network, and less stable allosteric
communication pathways in Lbc, compared to the nonconstitutively active
homologue, emerged as major determinants of the constitutive activity.
Independent of the state, the essential dynamics of the two RhoGEFs
is contributed by the last 10 amino acids of Dbl homology (DH) and
the whole pleckstrin homology (PH) domains and tends to be equalized
by the presence of RhoA. The catalytic activity of the RhoGEF relies
on the scaffolding action of the DH domain that primarily turns the
switch I (SWI) of RhoA on itself through highly conserved amino acids
participating in the stability core and essential for function. Changes
in the conformation of SWI and disorganization of the RhoA regions
deputed to nucleotide binding are among the major RhoGEF effects leading
to GDP release. Binding of RhoA reorganizes the allosteric communication
on RhoGEF, strengthening the communication among the canonical RhoA
binding site on DH, a secondary RhoA binding site on PH, and the binding
site for heterotrimeric G proteins, suggesting dual roles for RhoA
as a catalysis substrate and as a regulatory protein. The structure
network-based analysis tool employed in this study proved to be useful
for predicting potentially druggable regulatory sites in protein structures.