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Crystal structure of the archaeosine synthase QueF-like-Insights into amidino transfer and tRNA recognition by the tunnel fold.

ABSTRACT: The tunneling-fold (T-fold) structural superfamily has emerged as a versatile protein scaffold of diverse catalytic activities. This is especially evident in the pathways to the 7-deazaguanosine modified nucleosides of tRNA queuosine and archaeosine. Four members of the T-fold superfamily have been confirmed in these pathways and here we report the crystal structure of a fifth enzyme; the recently discovered amidinotransferase QueF-Like (QueF-L), responsible for the final step in the biosynthesis of archaeosine in the D-loop of tRNA in a subset of Crenarchaeota. QueF-L catalyzes the conversion of the nitrile group of the 7-cyano-7-deazaguanine (preQ0 ) base of preQ0 -modified tRNA to a formamidino group. The structure, determined in the presence of preQ0 , reveals a symmetric T-fold homodecamer of two head-to-head facing pentameric subunits, with 10 active sites at the inter-monomer interfaces. Bound preQ0 forms a stable covalent thioimide bond with a conserved active site cysteine similar to the intermediate previously observed in the nitrile reductase QueF. Despite distinct catalytic functions, phylogenetic distributions, and only 19% sequence identity, the two enzymes share a common preQ0 binding pocket, and likely a common mechanism of thioimide formation. However, due to tight twisting of its decamer, QueF-L lacks the NADPH binding site present in QueF. A large positively charged molecular surface and a docking model suggest simultaneous binding of multiple tRNA molecules and structure-specific recognition of the D-loop by a surface groove. The structure sheds light on the mechanism of nitrile amidation, and the evolution of diverse chemistries in a common fold. Proteins 2016; 85:103-116. © 2016 Wiley Periodicals, Inc.

SUBMITTER: Mei X

PROVIDER: S-EPMC5167649 | biostudies-literature | 2017 Jan

REPOSITORIES: biostudies-literature

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Crystal structure of the archaeosine synthase QueF-like-Insights into amidino transfer and tRNA recognition by the tunnel fold.

Mei Xianghan X Alvarez Jonathan J Bon Ramos Adriana A Samanta Uttamkumar U Iwata-Reuyl Dirk D Swairjo Manal A MA

Proteins 20161120 1

The tunneling-fold (T-fold) structural superfamily has emerged as a versatile protein scaffold of diverse catalytic activities. This is especially evident in the pathways to the 7-deazaguanosine modified nucleosides of tRNA queuosine and archaeosine. Four members of the T-fold superfamily have been confirmed in these pathways and here we report the crystal structure of a fifth enzyme; the recently discovered amidinotransferase QueF-Like (QueF-L), responsible for the final step in the biosynthesi ...[more]

PMID: 27802572

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Project description:Archaeosine (G+) is a structurally complex modified nucleoside found quasi-universally in the tRNA of Archaea and located at position 15 in the dihydrouridine loop, a site not modified in any tRNA outside the Archaea G+ is characterized by an unusual 7-deazaguanosine core structure with a formamidine group at the 7-position. The location of G+ at position 15, coupled with its novel molecular structure, led to a hypothesis that G+ stabilizes tRNA tertiary structure through several distinct mechanisms. To test whether G+ contributes to tRNA stability and define the biological role of G+, we investigated the consequences of introducing targeted mutations that disrupt the biosynthesis of G+ into the genome of the hyperthermophilic archaeon Thermococcus kodakarensis and the mesophilic archaeon Methanosarcina mazei, resulting in modification of the tRNA with the G+ precursor 7-cyano-7-deazaguansine (preQ0) (deletion of arcS) or no modification at position 15 (deletion of tgtA). Assays of tRNA stability from in vitro-prepared and enzymatically modified tRNA transcripts, as well as tRNA isolated from the T. kodakarensis mutant strains, demonstrate that G+ at position 15 imparts stability to tRNAs that varies depending on the overall modification state of the tRNA and the concentration of magnesium chloride and that when absent results in profound deficiencies in the thermophily of T. kodakarensis IMPORTANCE Archaeosine is ubiquitous in archaeal tRNA, where it is located at position 15. Based on its molecular structure, it was proposed to stabilize tRNA, and we show that loss of archaeosine in Thermococcus kodakarensis results in a strong temperature-sensitive phenotype, while there is no detectable phenotype when it is lost in Methanosarcina mazei Measurements of tRNA stability show that archaeosine stabilizes the tRNA structure but that this effect is much greater when it is present in otherwise unmodified tRNA transcripts than in the context of fully modified tRNA, suggesting that it may be especially important during the early stages of tRNA processing and maturation in thermophiles. Our results demonstrate how small changes in the stability of structural RNAs can be manifested in significant biological-fitness changes.

Dataset Information

Crystal structure of the archaeosine synthase QueF-like-Insights into amidino transfer and tRNA recognition by the tunnel fold.

Publications

Crystal structure of the archaeosine synthase QueF-like-Insights into amidino transfer and tRNA recognition by the tunnel fold.

Similar Datasets

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