Structural Insights into the Interaction of the Nip7 PUA Domain with Polyuridine RNA
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- 作者：Patrí ; cia P. Coltri ; Beatriz G. Guimarã ; es ; Daniela C. Granato ; Juliana S. Luz ; Elaine C. Teixeira ; Carla C. Oliveira ; Nilson I. T. Zanchin
- 刊名：Biochemistry
- 出版年：2007
- 出版时间：December 11, 2007
- 年：2007
- 卷：46
- 期：49
- 页码：14177 - 14187
- 全文大小：832K
- 年卷期：v.46,no.49(December 11, 2007)
- ISSN：1520-4995
文摘
The conserved protein Nip7 is involved in ribosome biogenesis, being required for proper27S pre-rRNA processing and 60S ribosome subunit assembly in Saccharomyces cerevisiae. Yeast Nip7pinteracts with nucleolar proteins and with the exosome subunit Rrp43p, but its molecular function remainsto be determined. Solution of the Pyrococcus abyssi Nip7 (PaNip7) crystal structure revealed a monomericprotein composed by two alpha-beta domains. The N-terminal domain is formed by a five-strandedantiparallel -sheet surrounded by three -helices and a 310 helix while the C-terminal, a mixed -sheetdomain composed by strands 8 to 12, one -helix, and a 310 helix, corresponds to the conserved PUAdomain (after Pseudo-Uridine synthases and Archaeosine-specific transglycosylases). By combiningstructural analyses and RNA interaction assays, we assessed the ability of both yeast and archaeal Nip7orthologues to interact with RNA. Structural alignment of the PaNip7 PUA domain with the RNA-interacting surface of the ArcTGT (archaeosine tRNA-guanine transglycosylase) PUA domain indicatedthat in the archaeal PUA domain positively charged residues (R151, R152, K155, and K158) are involvedin RNA interaction. However, equivalent positions are occupied by mostly hydrophobic residues (A/G160, I161, F164, and A167) in eukaryotic Nip7 orthologues. Both proteins can bind specifically topolyuridine, and RNA interaction requires specific residues of the PUA domain as determined by site-directed mutagenesis. This work provides experimental verification that the PUA domain mediates Nip7interaction with RNA and reveals that the preference for interaction with polyuridine sequences is conservedin Archaea and eukaryotic Nip7 proteins.