IBR   13079
INSTITUTO DE BIOLOGIA MOLECULAR Y CELULAR DE ROSARIO
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
RNA induced folding of the first double stranded RNA binding domain from A. thaliana DCL1
Autor/es:
BURDISSO PAULA; SUAREZ IRINA PAULA; HAILS GUILLERMO; BENOIT MATHIEW; BOISBOUVIER JEROME
Reunión:
Congreso; 18th ISMAR (International Society of Magnetic Resonance) Meeting; 2013
Resumen:
MicroRNAs are essential gene regulators in
multicellular organisms. Plant miRNAs are processed in the nucleus by a protein
complex formed by DICER-LIKE1 (DCL1), HYL1 and SERRATE. DCL1 has a central role
in the recognition and processing of the heterogeneous plant precursors. Little
is known about the structural aspects of this protein. In a previous work, we
characterized the second dsRBD of DCL1. Here we present a structural
characterization of the first DCL1 double stranded RNA binding domain.
In contrast with canonical dsRBDs, this domain binds
dsDNA with a similar affinity as dsRNA, the same as DCL1-dsRBD2.
We found that both
domains cooperate for substrate binding, but not for DNA binding.
Quite unexpectedly we
found that dsRBD1 is intrinsically disordered, even in the context of flanking
domains, but folds upon binding substrate RNA. We have assigned the backbone
resonances corresponding to the free unfolded and bound folded protein. The
free protein, though intrinsically disordered, shows a tendency to populate
folded conformations that correspond to those expected for a dsRBD. The
assignment of the bound form allowed us to calculate the fold of the protein in
complex with dsRNA employing CS-Rosetta. This analysis shows that this domain
acquires the dsRBD fold when bound to substrate. We further found that the
negatively charged surface provided by SDS micelles shifts the protein towards
folded conformations, but these are different from that found in the complex
with RNA. Experimental comparison with dsRBDs from other Dicer proteins show
that the intrinsically disorder nature is exclusive of DCL1-dsRBD1. Sequence
analysis and in vivo function of the protein suggests that in this domain the
instability of the free form and the ability to fold upon binding the substrate
have to be essential for the function of the whole DCL1 enzyme in miRNA
processing.