Circular dichroism and fluorescence studies on five mutant forms of protein synthesis initiation factor eIF‐4E, from the yeast Saccharomyces cerevisiae

FEBS Letters

Kay, Cyril M.²  Edery, Isaac¹  Altmann, Michael¹  Sonenberg, Nahum¹  McCubbin, William D.² 

[1] Medical Research Council of Canada Group in Protein Structure and Function, Department of Biochemistry, University of Alberta, Edmonton, Alberta T6G 2H7 Canada;Department of Biochemistry, McGill University, Montreal, PQ H3G 1Y6, Canada

关键词: Tryptophan;    CD;    Fluorescence spectroscopy;    Eukaryotic initiation factor-4E;    (S. cerevisiae);    eIF-4E;    eukaryotic initiation factor-4E;    K SV;    eff;    effective Stern-Volmer constant;    W n F;    tryptophan to phenylalanine mutant of eIF-4E at position n in the polypeptide chain;    m7GpppG;    7-methylguanosine-5′-triphosphate 5′-guanosine;   

DOI  :  10.1016/0014-5793(89)80233-9

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

CD studies have shown that five tryptophan to phenylalanine (W→F) mutants of eukaryotic initiation factor-4E (eIF-4E) contain low amounts of α-helix, the main elements of secondary structure being β-sheets/turns and aperiodic regions. Interactions with the cap analog m⁷GpppG are accompanied by changes in overall secondary structure which include reductions, and in one case an increase in α-helix content, as well as increases in total β-structure (3 mutant forms) and decreases in total β-structure (2 mutant forms). These changes may also involve more significant perturbations of localized regions containing phenylalanine residues either involved in nucleotide binding, or close to the nucleotide-binding site. Measurements of intrinsic Trp fluorescence have shown different quantum yields and reduced m⁷GpppG-induced quenching (with one exception). Acrylamide quenching studies yielded similar parameters for 4 of the mutants but 1 form displayed significantly reduced values. Melting experiments showed that the Trp fluorescence of 4 of the mutants decreased as the temperature was increased, this effect being reduced in 3 cases in the presence of m⁷GpppG. W 58 F showed an increase in fluorescence as the temperature was raised and this effect was accentuated in the presence of nucleotide. A preliminary attempt has been made to correlate the spectroscopic data with the known biological importance of the individual Trp residues.

【 授权许可】

Unknown