Bifunctional activity of deoxyhypusine synthase/hydroxylase from Trichomonas vaginalis

Quintas-Granados, Laura Itzel; Gamez, Bertha Isabel Carvajal; Villalpando, José Luis; Ortega‐López, Jaime; Arroyo, Rossana; Azuara-Licéaga, Elisa; Álvarez-Sánchez, María Elizbeth

doi:10.1016/j.biochi.2015.09.027

Cited by 9 publications

(6 citation statements)

References 57 publications

(87 reference statements)

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“…Therefore, the hypusination step might be needed to stabilize the modified form of the substrate. In any case, the absence of hypusinated aIF5A from the reaction products seems to exclude the possibility of aDHS being a bifunctional enzyme as described in T. vaginalis (Quintas-Granados et al 2016). The identification of the aIF5A interactome (Table 2) bears the potential to unravel the function(s) involved in hypusination of aIF5A.…”

Section: Discussionmentioning

confidence: 86%

“…However, the hypusination pathway remains obscure since no DOHH orthologous have been found in archaeal genomes. In Trichomonas vaginalis , hypusination of eIF5A is most likely the result of the catalytic activity of a single bifunctional enzyme (TvDHS), which performs both the DHS and DOHH reactions (Quintas-Granados et al 2016). Hence, we hypothesized that either Sso aDHS performs the synthesis of only deoxyhypusine, and then a second enzyme, still to be discovered, catalyzes hypusination or, alternatively, aDHS acts like the bifunctional TvDHS catalyzing both modifications.…”

Section: Resultsmentioning

confidence: 99%

“…The latter contains only deoxyhypusinylated aIF5A, whose synthesis differs from the canonical eukaryotic pathway: in the first reaction, the DHS enzyme transfers agmatine to the aIF5A lysine, while in the second reaction the agmatinase enzyme leads to production of deoxyhypusine. Alternatively, as described for T. vaginalis , the two modification reactions can be catalyzed by DHS, endowed with bifunctional activity (Quintas-Granados et al 2016). In any case, modification of that specific lysine seems to be important since at least some Archaea (e.g.…”

Section: Introductionmentioning

confidence: 99%

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Modification of translation factor aIF5A from Sulfolobus solfataricus

et al. 2018

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Eukaryotic eIF5A and its bacterial orthologue EF-P are translation elongation factors whose task is to rescue ribosomes from stalling during the synthesis of proteins bearing particular sequences such as polyproline stretches. Both proteins are characterized by unique post-translational modifications, hypusination and lysinylation, respectively, which are essential for their function. An orthologue is present in all Archaea but its function is poorly understood. Here, we show that aIF5A of the crenarchaeum Sulfolobus solfataricus is hypusinated and forms a stable complex with deoxyhypusine synthase, the first enzyme of the hypusination pathway. The recombinant enzyme is able to modify its substrate in vitro resulting in deoxyhypusinated aIF5A. Moreover, with the aim to identify the enzyme involved in the second modification step, i.e. hypusination, a set of proteins interacting with aIF5A was identified.Electronic supplementary materialThe online version of this article (10.1007/s00792-018-1037-4) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 86%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Modification of translation factor aIF5A from Sulfolobus solfataricus

et al. 2018

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“…Both enzymes are solely dedicated to the hypusination of eIF5A and are essential in most eukaryotes. In this sense, while DHPS is essential in archaea, a DOHH ortholog has not been found, with DHPS having both enzymatic activities in some cases [ 98 , 99 ]. As the deoxyhypusine formation by DHPS is reversible in vitro, it has been proposed that the hydroxylation step could have evolved as a stabilization mechanism for the deoxyhypusine modification [ 100 , 101 ].…”

Section: The Relevance Of Iron In Translation Elongationmentioning

confidence: 99%

Iron in Translation: From the Beginning to the End

2021

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Iron is an essential element for all eukaryotes, since it acts as a cofactor for many enzymes involved in basic cellular functions, including translation. While the mammalian iron-regulatory protein/iron-responsive element (IRP/IRE) system arose as one of the first examples of translational regulation in higher eukaryotes, little is known about the contribution of iron itself to the different stages of eukaryotic translation. In the yeast Saccharomyces cerevisiae, iron deficiency provokes a global impairment of translation at the initiation step, which is mediated by the Gcn2-eIF2α pathway, while the post-transcriptional regulator Cth2 specifically represses the translation of a subgroup of iron-related transcripts. In addition, several steps of the translation process depend on iron-containing enzymes, including particular modifications of translation elongation factors and transfer RNAs (tRNAs), and translation termination by the ATP-binding cassette family member Rli1 (ABCE1 in humans) and the prolyl hydroxylase Tpa1. The influence of these modifications and their correlation with codon bias in the dynamic control of protein biosynthesis, mainly in response to stress, is emerging as an interesting focus of research. Taking S. cerevisiae as a model, we hereby discuss the relevance of iron in the control of global and specific translation steps.

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“…For DOHH no homolog has been identified yet (Park, 2006;Wolff et al, 2007). In some higher eukaryote, a DHS was described to be bifunctional, performing the classic DHS-reaction as well as the DOHH catalyzed dehydration reaction (Quintas-Granados et al, 2016). For the studied archaea, however, such bifunctionality of DHS was not observed (Bassani et al, 2018).…”

Section: Post-translation Modifications Of E/aif5amentioning

confidence: 94%

Recognition Elements for Elongation Factor P on the Ribosome

Frister¹

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Translation elongation is a multi-step process orchestrated by elongation factors.Elongation factors G and Tu are required for each round of translation elongation, whereas elongation factor P is only required to assist the translation of poly(Pro) sequences. Upon incorporation of consecutive proline residues the ribosome is stalled; this stalling is alleviated by EF-P. While the catalytic mechanism of EF-P is well described, the determinants of EF-P binding remain unknown. Structural data and biochemical studies suggest the E-site codon, the peptidyl-tRNA, the ribosomal protein L1 and the post-translational modification of EF-P as key interaction partners during binding and EF-P assisted catalysis.In this thesis we developed a FRET based EF-P binding assay using fluorescence-labeled ribosome complexes and a quencher-labeled EF-P. We combined the binding assay with different EF-P activity assays to determine the contribution of each of the proposed interactions to the binding and the catalytic activity of EF-P. We found that EF-P binds to different ribosome complexes with similar rates. EF-P has a short residence time on complexes without poly(Pro) stalling sequences, which is significantly increased on poly(Pro)-stalled complexes. This high affinity state depends on the presence of several recognition elements in poly(Pro)-stalled complexes, in particular tRNA Pro in the P site and the polypeptide chain containing several sequential proline residues. The contextindependent association rates and the determined cellular concentration of EF-P suggest that the sampling of ribosome complexes by EF-P is kinetically controlled by the availability of a vacant E site. However, only poly(Pro)-stalled ribosome complexes provide the interactions required for the high-affinity binding of EF-P. The dissociation rates from Pro-stalled and non-stalled complexes match the reported rates of EF-P-assisted peptide bond formation. This suggests a mechanism in which the prolonged residence time for stalled complexes allows EF-P to position the peptidyl-tRNA in a catalytically active conformation and thereby to alleviate the stalling. After peptide bond formation the complex returns to the low affinity state, inducing dissociation of EF-P. The proposed kinetic regime allows EF-P to efficiently sample ribosomes with empty E sites, to recognize Pro-stalled complexes with high turnover rates and to alleviate stalling in a single functional cycle. Thus, our work demonstrates that the recruitment of EF-P is kinetically controlled contributing to a harmonized rate of translation. when fMet-tRNA fMet recognizes the AUG start codon displayed by the mRNA (Milon et al., 2008). Joining of the 50S subunit triggers the GTPase activity of IF2, causing GTP hydrolysis and resulting in the dissociation of IF1 and IF2. The dissociation of IF3 marks the formation of the translation competent 70S IC (Goyal et al., 2015;Grigoriadou et al., 2007;Tomsic et al., 2000). The translation competent 70S IC provides three binding sites for tRNAs, the acceptor site...

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Bifunctional activity of deoxyhypusine synthase/hydroxylase from Trichomonas vaginalis

Cited by 9 publications

References 57 publications

Modification of translation factor aIF5A from Sulfolobus solfataricus

Modification of translation factor aIF5A from Sulfolobus solfataricus

Iron in Translation: From the Beginning to the End

Recognition Elements for Elongation Factor P on the Ribosome

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