Yeast as a tool to express sugar acid transporters with biotechnological interest

Ribas, David Manuel Nogueira; Sá‐Pessoa, Joana; Soares-Silva, Isabel; Paiva, Sandra; Nygård, Yvonne; Ruohonen, Laura; Penttilä, Merja; Casal, Margarida

doi:10.1093/femsyr/fox005

Cited by 12 publications

(15 citation statements)

References 51 publications

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“…Since SatP three-dimensional modelling obtained the best score for protein structure prediction, it was further considered for molecular docking analysis. Molecular docking simulations were performed as described before (Ribas et al, 2017).…”

Section: Three-dimensional Modelling and Molecular Dockingmentioning

confidence: 99%

“…for three independent experiments (n = 9). membrane carboxylate mediated transport under the conditions tested (Queirós et al, 2007;Ribas et al, 2017;Soares-Silva et al, 2007;Soares-Silva et al, 2015). Mid-exponential S. cerevisiae cells expressing GPR1 grown on glucose (minimal medium YNB-Glu) were collected, washed and incubated for 6 h in minimal medium containing 0.5% acetic acid (pH 6.0), as sole carbon and energy source.…”

Section: Gpr1 Mediates Acetate Transport In S Cerevisiaementioning

confidence: 99%

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The acetate uptake transporter family motif “NPAPLGL(M/S)” is essential for substrate uptake

Ribas

Soares-Silva

Vieira

et al. 2019

Fungal Genetics and Biology

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Organic acids are recognized as one of the most prevalent compounds in ecosystems, thus the transport and assimilation of these molecules represent an adaptive advantage for organisms. The AceTr family members are associated with the active transport of organic acids, namely acetate and succinate. The phylogenetic analysis shows this family is dispersed in the tree of life. However, in eukaryotes, it is almost limited to microbes, though reaching a prevalence close to 100% in fungi, with an essential role in spore development. Aiming at deepening the knowledge in this family, we studied the acetate permease AceP from Methanosarcina acetivorans, as the first functionally characterized archaeal member of this family. Furthermore, we demonstrate that the yeast Gpr1 from Yarrowia lipolytica is an acetate permease, whereas the Ady2 closest homologue in Saccharomyces cerevisiae, Fun34, has no role in acetate uptake. In this work, we describe the functional role of the AceTr conserved motif NPAPLGL(M/S). We further unveiled the role of the amino acid residues R122 and Q125 of SatP as essential for protein activity.

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Section: Three-dimensional Modelling and Molecular Dockingmentioning

confidence: 99%

Section: Gpr1 Mediates Acetate Transport In S Cerevisiaementioning

confidence: 99%

The acetate uptake transporter family motif “NPAPLGL(M/S)” is essential for substrate uptake

Ribas

Soares-Silva

Vieira

et al. 2019

Fungal Genetics and Biology

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“…By selecting SatP_Ck crystal structure as a template and using the Modeller homology modelling program [33] , the Ato1 3D structure was obtained. Visual analyses of Ato1 3D structure were performed using the Free Maestro version molecular modelling software as previously reported [34] .…”

Section: Methodsmentioning

confidence: 99%

New insights into the acetate uptake transporter (AceTr) family: Unveiling amino acid residues critical for specificity and activity

Rendulić

Alves

Azevedo-Silva

et al. 2021

Computational and Structural Biotechnology Journal

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“…The uptake of labeled carboxylic acids was assessed as previously described by Ribas et al (2017), using [1-14 C] acetic acid (Perkin Elmer, Waltham, MA) and [U-14 C] l-lactic acid (Perkin Helmer) with a specific activity of 2000 dpm/nmol. The data shown are mean values of at least three independent experiments.…”

Section: Transport Assaysmentioning

confidence: 99%

Evolutionary engineering reveals amino acid substitutions in Ato2 and Ato3 that allow improved growth ofSaccharomyces cerevisiaeon lactic acid

Baldi

Valk

Sousa-Silva

et al. 2021

FEMS Yeast Research

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In Saccharomyces cerevisiae, the complete set of proteins involved in transport of lactic acid across the cell membrane has not been determined. In this study we aimed to identify transport proteins not previously described to be involved in lactic acid transport via a combination of directed evolution, whole-genome resequencing and reverse engineering. Evolution of a strain lacking all known lactic acid transporters on lactate led to the discovery of mutated Ato2 and Ato3 as two novel lactic acid transport proteins. When compared to previously identified S. cerevisiae genes involved in lactic acid transport, expression of ATO3T284C was able to facilitate the highest growth rate (0.15 ± 0.01 h−1) on this carbon source. A comparison between (evolved) sequences and 3D models of the transport proteins showed that most of the identified mutations resulted in a widening of the narrowest hydrophobic constriction of the anion channel. We hypothesize that this observation, sometimes in combination with an increased binding affinity of lactic acid to the sites adjacent to this constriction, are responsible for the improved lactic acid transport in the evolved proteins.

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Yeast as a tool to express sugar acid transporters with biotechnological interest

Cited by 12 publications

References 51 publications

The acetate uptake transporter family motif “NPAPLGL(M/S)” is essential for substrate uptake

The acetate uptake transporter family motif “NPAPLGL(M/S)” is essential for substrate uptake

New insights into the acetate uptake transporter (AceTr) family: Unveiling amino acid residues critical for specificity and activity

Evolutionary engineering reveals amino acid substitutions in Ato2 and Ato3 that allow improved growth ofSaccharomyces cerevisiaeon lactic acid

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