J Mol Biol 2005 Apr 29;3482:265-79. doi: 10.1016/j.jmb.2005.02.058.

Solution structure and backbone dynamics of the KH-QUA2 region of the Xenopus STAR/GSG quaking protein.

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The Quaking protein belongs to the family of STAR/GSG domain RNA-binding proteins and is involved in multiple cell signalling and developmental processes in vertebrates, including the formation of myelin. Heteronuclear NMR methods were used to determine the solution structure of a 134 residue fragment spanning the KH and QUA2 homology regions of the Quaking protein from Xenopus laevis (pXqua) in the absence of RNA. The protein is shown to adopt an extended type I KH domain fold that is connected to a structured alpha-helix in the C-terminal QUA2 region by means of a highly flexible linker. A comparison with the solution structure of the related protein splicing factor 1 (SF1) indicates that most aspects of the RNA-binding interface are conserved in pXqua, although the "variable loop" region that follows the second beta-strand possesses two additional alpha-helices. The structure of pXqua provides an appropriate template for building models of important homologues, such as GLD-1 and Sam68. Measurements of the (15)N relaxation parameters of pXqua confirm that the polypeptide backbone of the QUA2 region is more dynamic than that of the KH portion, and that the C-terminal helix is partially structured in the absence of RNA. By comparison with a random coil reference state, the nascent structure in the QUA2 region is estimated to contribute 15.5kJmol(-1) to the change in conformational free energy that occurs on forming a complex with RNA. Since STAR/GSG proteins may regulate alternative splicing by competing with SF1 in the nucleus for specific branch-point sequences that signal intronic RNA, the formation of secondary structure in the QUA2 region in the unbound state of pXqua has important functional consequences.

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Species referenced: Xenopus laevis
Genes referenced: khdrbs1 qki sf1