De Rocquigny H, Ficheux D, Gabus C, Allain B, Fournie-Zaluski M C, Darlix J L, Roques B P
Unite de Pharmacochimie moléculaire et structurale, U266 INSERM, URA D1500 CNRS, Université René Descartes, Paris, France.
Nucleic Acids Res. 1993 Feb 25;21(4):823-9. doi: 10.1093/nar/21.4.823.
The 56 amino acid nucleocapsid protein (NCp10) of Moloney Murine Leukemia Virus, contains a CysX2CysX4HisX4Cys zinc finger flanked by basic residues. In vitro NCp10 promotes genomic RNA dimerization, a process most probably linked to genomic RNA packaging, and replication primer tRNA(Pro) annealing to the initiation site of reverse transcription. To characterize the amino-acid sequences involved in the various functions of NCp10, we have synthesized by solid phase method the native protein and a series of derived peptides shortened at the N- or C-terminus with or without the zinc finger domain. In the latter case, the two parts of the protein were linked by a Glycine - Glycine spacer. The in vitro studies of these peptides show that nucleic acid annealing activities of NCp10 do not require a zinc finger but are critically dependent on the presence of specific sequences located on each side of the CCHC domain and containing proline and basic residues. Thus, deletion of 11R or 49PRPQT, of the fully active 29 residue peptide 11RQGGERRRSQLDRDGGKKPRGPRGPRPQT53 leads to a complete loss of NCp10 activity. Therefore it is proposed that in NCp10, the zinc finger directs the spatial recognition of the target RNAs by the basic domains surrounding the zinc finger.
莫洛尼鼠白血病病毒的56个氨基酸的核衣壳蛋白(NCp10)含有一个由碱性残基侧翼的CysX2CysX4HisX4Cys锌指结构。在体外,NCp10促进基因组RNA二聚化,这一过程很可能与基因组RNA包装以及复制引物tRNA(Pro)退火至逆转录起始位点有关。为了表征参与NCp10各种功能的氨基酸序列,我们通过固相法合成了天然蛋白以及一系列在N端或C端缩短的衍生肽,这些肽带有或不带有锌指结构域。在后一种情况下,蛋白质的两部分通过甘氨酸-甘氨酸间隔区连接。对这些肽的体外研究表明,NCp10的核酸退火活性不需要锌指结构,但严重依赖于位于CCHC结构域两侧且含有脯氨酸和碱性残基的特定序列的存在。因此,完全活性的29个残基肽11RQGGERRRSQLDRDGGKKPRGPRGPRPQT53中11R或49PRPQT的缺失会导致NCp10活性完全丧失。因此,有人提出在NCp10中,锌指通过锌指周围的碱性结构域指导对靶RNA的空间识别。