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. 2011 Mar 8;50(9):1429-31.
doi: 10.1021/bi102057m. Epub 2011 Feb 2.

Large favorable enthalpy changes drive specific RNA recognition by RNA recognition motif proteins

Krystle J McLaughlin  1 , Jermaine L Jenkins, Clara L Kielkopf
Affiliations

Large favorable enthalpy changes drive specific RNA recognition by RNA recognition motif proteins

Krystle J McLaughlin et al. Biochemistry. .

Abstract

The RNA recognition motif (RRM) is a prevalent class of RNA binding domains. Although a number of RRM/RNA structures have been determined, thermodynamic analyses are relatively uncommon. Here, we use isothermal titration calorimetry to characterize single-stranded (ss)RNA binding by four representative RRM-containing proteins: (i) U2AF(65), (ii) SXL, (iii) TIA-1, and (iv) PAB. In all cases, ssRNA binding is accompanied by remarkably large favorable enthalpy changes (-30 to -60 kcal mol(-1)) and unfavorable entropy changes. Alterations of key RRM residues and binding sites indicate that under the nearly physiological conditions of these studies, large thermodynamic changes represent a signature of specific ssRNA recognition by RRMs.

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Figures

Figure 1
Figure 1
Structures of the second RRMs of (A) SXL (PDB ID 1B7F) or (B) PAB (PDB ID 1CVJ). RNP residues altered by mutagenesis are colored yellow. (C) U2AF65, TIA-1, SXL, and PAB domain organization and constructs used in this study. RRM, RNA recognition motif; RS, arginine-serine rich domain; ULM, UHM ligand motif; UHM, U2AF homology motif; Gly-rich, glycine-rich domain; Pro-rich, proline-rich domain; Gln-rich, glutamine-rich domain; PABC, peptide-binding domain.
Figure 2
Figure 2
Representative isotherms for titration of (A) U20 RNA into SXL RRM12; (B) A20 RNA into SXL RRM12; (C) U20 RNA into SXL RRM12Mut in 100 mM NaCl, 25 mM BES pH 7.4 at 303 K. All other isotherms are represented in the Supplementary Data. (D) Bar graph representations of thermodynamic changes. Left panel, specific RNA binding by wild-type RRM-containing proteins: U2AF65RRM12U/U20, SXL RRM12/U20, TIA-1 RRM23/U20, and PAB RRM12/A20. (*)Averaged nonredundant enthalpy (www.bioinfodatabase.com/pint/) and entropy (13) changes for protein-protein interactions. Central panel, noncognate RNA binding: SXL RRM12/A20 and PAB RRM12/U20. Right panel, RNP mutant proteins: SXL RRM12Mut/U20 and PAB RRM12Mut/A20.
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