Conserved linker length in double dsRBD proteins from plants restricts interdomain motion

LEANDRO C. TABARES; RASIA, RODOLFO M.; MASCALI, FLORENCIA C.

La Plata

Congreso; XLVII Reunión Anual de la Sociedad Argentina de Biofísica; 2018

Sociedad Argentina de Biofisica

Multidomain proteins are ubiquitous in nature. Domain shuffling and duplication have helped organisms to achieve a wide range of functions by combining a limited number of individual modules. The domains in multidomain proteins are connected by sequences of aminoacids of varying length and rigidity. HYL1 is a protein from Arabidopsis thaliana that participates in microRNA biogenesis in plants whose precise function is not known at present. Analysis of HYL1 sequence showed a domain architecture consisting of two double stranded RNA binding domains (dsRBDs) connected by a 17-residue linker and followed by a long unstructured C-terminal dispensable for its function in vivo. In the present work we studied how the linker restricts the mobility between both domains. Inter domain distances measured by PELDOR on double labeled protein samples show a narrow distribution around 4 nm. NMR Paramagnetic Relaxation Enhancement (PRE) on single labeled protein samples shows an asymmetry in transient domain-domain interactions. By means of ensemble simulations and calculation of PREs we found that the domains explore a restricted conformational space. Sequence analysis of linkers between double dsRBD plant proteins shows a high conservation in linker length. Altogether our results put forward the functional importance of the linkers between dsRBD modules in multidomain proteins.