STRUCTURE AND DYNAMICS OF THE RRM1 DOMAIN OF THE POST-TRANSCRIPTIONAL REGULATOR HUR: IMPLICATIONS FOR RNA RECOGNITION
Mujo, A.1, Carneiro, L. A. M.2, Anobom, C. D. and Pinheiro, A. S.1
1 Departamento de Bioquímica, Instituto de Química, Universidade Federal do Rio de Janeiro, Rio de Janeiro-RJ, Brasil.
2 Departamento de Imunologia, Instituto de Microbiologia Paulo de Góes, Universidade Federal do Rio de Janeiro, Rio de Janeiro-RJ, Brasil.
Introduction: HuR is a ubiquitous protein that recognizes AREs in mRNA, thereby interfering with the fate of protein translation. This protein plays a central role on the outcome of the inflammatory response as it may stabilize or silence the mRNAs of key components of the immune system. HuR is able to interact with other RBPs, reflecting a complex network that dictates mRNAs post-transcriptional control. HuR is composed of three functional domains, known as RRM1, RRM2 and RRM3. RRM1 is the major mRNA recognition domain of HuR. Here, we report the structure and dynamics of RRM1 determined by NMR spectroscopy. Materials and Methods: NMR spectra were collected on a 15N- and a 15N/13C-labeled purified RRM1 sample using a Bruker 800 MHz spectrometer. Sequence-specific resonance assignment and protein structure determination were performed using programs cara and CCPN/Aria, respectively. Backbone dynamics were derived from 15N-relaxation measurements including R1, R2 and 15N[1H] NOE. Results: RRM1’s overall fold consists of two a-helices and a four-stranded b-sheet, with a b1-a1-b2-b3-a2-b4 topology and a b-hairpin between a2 and b4. Residues Asn21, Asn26, Tyr27, Gln29, Lys50, Phe65, Lys89, Thr90, Lys92, Arg97 and Ser99 are directly involved in mRNA binding. Despite the first 18 residues, which are unstructured in solution, high flexibility of the protein backbone is observed for three regions of RRM1, including residues 28-30, 55-57 and 88-90. In addition, residues 29-32 and 55-60 show backbone internal motions on the ms-ms time scale. Conclusion: The dynamic properties observed for the RRM1 domain of HuR, especially for residues involved in RNA binding, may indicate a correlation between structural plasticity and the recognition of specific RNA elements.
Financial Support:FAPERJ and CNPq. Keywords: HuR, RNA, structure, dynamics, NMR.
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