A characteristic arrangement of aromatic amino acid residues in the solution structure of the amino-terminal RNA-binding domain of Drosophila sex-lethal.
The Sex-lethal (Sxl) protein from Drosophila melanogaster has two RNA-binding domains (RBDs). As the amino-terminal RBD (RBD1) of the Sxl protein exhibits low sequence homology to the typical RBDs, particularly at the putative functional residues, it was difficult to unambiguously locate the RNP1 and RNP2 motifs. Therefore, in the present study, we defined the amino and carboxy-terminal borders of the first RNA-binding domain (RBD1) of the Sxl protein by limited tryptic digestion. By replacement of Phe166 by Tyr, we constructed a highly soluble mutant, which exhibits the same RNA-binding properties as those of the wild-type. Using this mutant protein, we performed NMR measurements, and elucidated the secondary and tertiary structures of the Sxl RBD1 in solution. The betaalphabetabetaalphabeta folding pattern is conserved in the solution structure of the Sxl RBD1, as in other reported RBD structures. This allowed us to identify both the RNP1 and RNP2 motifs of the Sxl RBD1 unambiguously. Intriguingly, the RNP2 motif of the Sxl RBD1 has an Ile residue at the second position, which is generally occupied by an aromatic amino acid residue in RBDs and has been suggested to be involved in their RNA binding. Furthermore, the loop region between beta2 and beta3 of the Sxl RBD1 has an exceptional cluster of aromatic amino acid residues, in place of the normal basic amino acid cluster. In contrast, the second RBD of Sxl does not exhibit these characteristic features.
Inoue
M
M
Department of Biophysics and Biochemistry, Graduate School of Science, The University of Tokyo, Tokyo, Hongo, Bunkyo-ku, 113, Japan.
Muto
Y
Y
Sakamoto
H
H
Kigawa
T
T
Takio
K
K
Shimura
Y
Y
Yokoyama
S
S
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D029721:Drosophila Proteins; D004331:Drosophila melanogaster / Q000737:chemistry; D004926:Escherichia coli; D007301:Insect Hormones / Q000737:chemistry*; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D016601:RNA-Binding Proteins / Q000737:chemistry*; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D016415:Sequence Alignment; D012995:Solubility; D012996:Solutions
D029721:Drosophila Proteins; D007301:Insect Hormones; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D012996:Solutions; C067777:Sxl protein, Drosophila