The N-terminal domain of the Drosophila histone mRNA binding protein, SLBP, is intrinsically disordered with nascent helical structure.
Stem-loop binding protein (SLBP) is a 31 kDa protein that is central to the regulation of histone mRNAs and is highly conserved in metazoans. In vertebrates, the N-terminal domain of SLBP has sequence determinants necessary for histone mRNA translation, SLBP degradation, cyclin binding, and histone mRNA import. We have used high-resolution NMR spectroscopy and circular dichroism to characterize the structural and dynamic features of this domain of SLBP from Drosophila (dSLBP). We report that the N-terminal domain of dSLBP is stably unfolded but has nascent helical structure at physiological pH and native-like solution conditions. The conformational and dynamic properties of the isolated domain are mimicked in a longer 175-residue region of the N-terminus, as well as in the full-length protein. Complete resonance assignments, secondary structure propensity, and motional properties of a 91-residue N-terminal domain (G17-K108) of dSLBP are reported here. The deviation of (1)H(alpha), (13)C(alpha), and (13)C(beta) chemical shifts from random coil reveals that there are four regions between residues I28-A45, S50-L57, S66-G75, and F91-N96 that have helical propensity. These regions also have small but positive heteronuclear NOEs, interresidue d(NN) NOEs, and small but significant protection from solvent exchange. However the lack of medium- and long-range NOEs in 3D (15)N- and (13)C-edited spectra, fast amide proton exchange rates (all greater than 1 s(-1)), and long (15)N relaxation (T(1), T(2)) times suggest that the domain from dSLBP does not adopt a well-defined tertiary fold. The backbone residual dipolar couplings (RDCs) for this domain are small and lie close to 0 Hz (+/-2 Hz) for most residues with no well-defined periodicity. The implications of this unfolded state for the function of dSLBP in regulating histone metabolism are discussed.
Thapar
Roopa
R
Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, North Carolina 27599, USA. thapar@email.unc.edu
Mueller
Geoffrey A
GA
Marzluff
William F
WF
D000595:Amino Acid Sequence; D000818:Animals; D002942:Circular Dichroism; D004330:Drosophila / Q000737:chemistry*; D029721:Drosophila Proteins / Q000737:chemistry / Q000378:metabolism*; D006657:Histones / Q000235:genetics*; D008969:Molecular Sequence Data; D011487:Protein Conformation; D012333:RNA, Messenger / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*
D029721:Drosophila Proteins; D006657:Histones; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; C490155:SLBP protein, Drosophila