Investigation of zinc-binding affinities of moloney murine leukemia virus nucleocapsid protein and its related zinc finger and modified peptides
✍ Scribed by Yves Mely; Fabrice Cornllle; Marie-Claude Fournié-Zaluski; Jean-Luc Darlix; Bernard F. Roques; Dominique Gérard
- Publisher
- Wiley (John Wiley & Sons)
- Year
- 1991
- Tongue
- English
- Weight
- 669 KB
- Volume
- 31
- Category
- Article
- ISSN
- 0006-3525
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✦ Synopsis
Nucleocapsid proteins of retroviruses are small basic, nucleic acid-binding proteins with either one or two "Cys-His" boxes, which have been shown to be involved in genomic RNA dimerization, encapsidation, and replication primer tRNA annealing to the initiation site for reverse transcription. The nucleocapsid ( NC ) protein of Moloney murine leukemia virus (MoMuLV NCplO) is made up of 56 residues with one Cys-His motif. The Zn'+binding affinities and induced conformational changes of NCplO were investigated by following the fluorescence of Trp 35 located in the Cys-His domain. At pH 7.5, NCplO was shown to bind Zn2+ at a 1 : 1 ratio with a very high apparent binding constant of 1.2 (k0.3) 1013M-'. A similar apparent binding constant was obtained for a 19-residue peptide encompassing the Cys-His box, designated the "zinc finger motif," indicating that it contains most if not all the information to bind Zn2+ tightly. Changing Trp 35 to Phe in the peptide did not affect the Zn2+ affinity, indicating that Trp 35 is not crucial for Zn2+ binding. On the contrary, replacing Cys 29 by Ser, the chemical modification or oxidation of the three Cys sharply reduced Zn2+ affinity, confirming the essential role of Cys in Zn2+ binding. In addition, fluorescence and energy transfer data suggested that Zn 2+ binding modifies the Trp 35 environment but not its solvent exposure, and increases the average distance between Tyr 28 and Trp 35 by about 2 A. These data suggest that Zn2' binding to retroviral NC protein is biologically relevant.