Abstract
BackgroundGlobal climate change and fossil fuels limitations have boosted the demand for robust and efficient microbial factories for the manufacturing of bio-based products from renewable feedstocks. In this regard, efforts have been done to enhance the enzyme-secreting ability of lignocellulose-degrading fungi, aiming to improve protein yields while taking advantage of their ability to use lignocellulosic feedstocks. Access to sugars in complex polysaccharides depends not only on their release by specific hydrolytic enzymes, but also on the presence of transporters capable of effectively transporting the constituent sugars into the cell. This study aims to identify and characterize xylose transporters from Aspergillus niger and Trichoderma reesei, two fungi that have been industrially exploited for decades for the production of lignocellulose-degrading hydrolytic enzymes.ResultsA hidden Markov model for the identification of xylose transporters was developed and used to analyze the A. niger and T. reesei in silico proteomes, yielding a list of candidate xylose transporters. From this list, three A. niger (XltA, XltB and XltC) and three T. reesei (Str1, Str2 and Str3) transporters were selected, functionally validated and biochemically characterized through their expression in a Saccharomyces cerevisiae hexose transport null mutant, engineered to be able to metabolize xylose but unable to transport this sugar. All six transporters were able to support growth of the engineered yeast on xylose but varied in affinities and efficiencies in the uptake of the pentose. Amino acid sequence analysis of the selected transporters showed the presence of specific residues and motifs recently associated to xylose transporters. Transcriptional analysis of A. niger and T. reesei showed that XltA and Str1 were specifically induced by xylose and dependent on the XlnR/Xyr1 regulators, signifying a biological role for these transporters in xylose utilization.ConclusionsThis study revealed the existence of a variety of xylose transporters in the cell factories A. niger and T. reesei. The particular substrate specificity and biochemical properties displayed by A. niger XltA and XltB suggested a possible biological role for these transporters in xylose uptake. New insights were also gained into the molecular mechanisms regulating the pentose utilization, at inducer uptake level, in these fungi. Analysis of the A. niger and T. reesei predicted transportome with the newly developed hidden Markov model showed to be an efficient approach for the identification of new xylose transporting proteins.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0564-4) contains supplementary material, which is available to authorized users.
Highlights
Global climate change and fossil fuels limitations have boosted the demand for robust and efficient microbial factories for the manufacturing of bio-based products from renewable feedstocks
The hidden Markov model specific for xylose transporters (HMMxylT) built from these sequences was used to mine the in silico proteomes of A. niger and T. reesei for putative xylose transporters
The A. niger HMMxylT output was compared with different transcriptome analysis studies where A. niger was grown in the presence of xylose, xylose–arabinose mixes, sugarcane bagasse, straw or willow [29,30,31,32,33]
Summary
Global climate change and fossil fuels limitations have boosted the demand for robust and efficient microbial factories for the manufacturing of bio-based products from renewable feedstocks In this regard, efforts have been done to enhance the enzyme-secreting ability of lignocellulose-degrading fungi, aiming to improve protein yields while taking advantage of their ability to use lignocellulosic feedstocks. Fossil fuels limitations and breakthroughs in the advanced biofuels market have boosted the demand for robust and efficient microbial cell factories In this regard, to enhance the enzyme-secreting ability of these fungi, efforts at multiple levels have been done: from studies at system level to understand the secretory process, to the use of more applied strategies focused on the improvement of protein yields [5, 6]. We estimate that for T. reesei 235 proteins can be classified as MFS transporters of which 113 can be classified as SP
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