The Overexpression of YALI0B07117g Results in EnhancedErythritol Synthesis from Glycerol by the Yeast Yarrowia lipolytica

The unconventional yeast Yarrowia lipolytica is used to produce erythritol from glycerol. In this study, the role of the erythrose reductase (ER) homolog YALI0B07117g in erythritol synthesis was analyzed. The deletion of the gene resulted in an increased production of mannitol (308%) and arabitol (204%) before the utilization of these polyols began. The strain overexpressing the YALI0B07117g gene was used to increase the erythritol yield from glycerol as a sole carbon source in batch cultures, resulting in a yield of 0.4 g/g. The specific consumption rate (qs) increased from 5.83 g/g/L for the WT strain to 8.49 g/g/L for the modified strain and the productivity of erythritol increased from 0.28 g/(L h) for the A101 strain to 0.41 g/(L h ) for the modified strain. The application of the research may prove positive for shortening the cultivation time due to the increased rate of consumption of the substrate combined with the increased parameters of erythritol synthesis.

Establishment of a near-contiguous genome sequence of the citric acid producing yeast Yarrowia lipolytica DSM 3286 with resolution of rDNA clusters and telomeres

Yarrowia lipolytica is an oleaginous yeast that is particularly suitable for the sustainable production of secondary metabolites. The genome of this yeast is characterized by its relatively large size and its high number of different rDNA clusters located in its telomeric regions. However, due to the presence of long repetitive elements in the sub-telomeric regions, rDNA clusters and telomeres are missing in current genome assemblies of Y. lipolytica. Here, we present the near-contiguous genome sequence of the biotechnologically relevant strain DSM 3286. We employed a hybrid assembly strategy combining Illumina and nanopore sequencing reads to integrate all six rDNA clusters as well as telomeric repeats into the genome sequence. By fine-tuning of DNA isolation and library preparation protocols, we were able to create ultra-long reads that not only contained multiples of mitochondrial genomes but also shed light on the inter- and intra-chromosomal diversity of rDNA cluster types. We show that there are ten different rDNA units present in this strain that additionally appear in a predefined order in a cluster. Based on single reads, we also demonstrate that the number of rDNA repeats in a specific cluster varies from cell to cell within a population.

Microbial Biosynthesis of Lactones: Gaps and Opportunities towards Sustainable Production

Lactones are volatile organic compounds widely present in foods. These chemicals are applied as flavors and fragrances in the food, cosmetics and pharmaceutical industries. Recently, the potential of lactones as green solvents and fuel precursors reinforced their role as platform compounds of future bio-based economies. However, their current mode of production needs to change. Lactones are mainly obtained through chemical synthesis or microbial biotransformation of hydroxy fatty acids. The latter approach is preferred but still needs to use more sustainable substrates. Hydroxy fatty acids are non-abundant and non-sustainable substrates from environmental, health and economic points of view. Therefore, it is urgent to identify and engineer microorganisms with the rare ability to biosynthesize lactones from carbohydrates or renewable lipids. Here, we firstly address the variety and importance of lactones. Then, the current understanding of the biosynthetic pathways involved in lactone biosynthesis is presented, making use of the knowledge acquired in microorganisms and fruits. From there, we present and make the distinction between biotransformation processes and de novo biosynthesis of lactones. Finally, the opportunities and challenges towards more sustainable production in addition to the relevance of two well-known industrial microbes, the filamentous fungus Ashbya gossypii and the yeast Yarrowia lipolytica, are discussed.

Bioproducts generation from carboxylate platforms by the non-conventional yeast Yarrowia lipolytica

In recent years, there has been a growing interest in the use of renewable sources for bio-based production aiming at developing sustainable and feasible approaches towards a circular economy. Among these renewable sources, organic wastes (OWs) can be anaerobically digested to generate carboxylates like volatile fatty acids (VFAs), lactic acid, and longer-chain fatty acids that are regarded as novel building blocks for the synthesis of value-added compounds by yeasts. This review discusses on the processes that can be used to create valuable molecules from OW-derived VFAs; the pathways employed by the oleaginous yeast Yarrowia lipolytica to directly metabolize such molecules; and the relationship between OW composition, anaerobic digestion, and VFA profiles. The review also summarizes the current knowledge about VFA toxicity, the pathways by which VFAs are metabolized, and the metabolic engineering strategies that can be employed in Y. lipolytica to produce value-added biobased compounds from VFAs.

The Role of Hexokinase and Hexose Transporters in Preferential Use of Glucose over Fructose and Downstream Metabolic Pathways in the Yeast Yarrowia lipolytica

The development of efficient bioprocesses requires inexpensive and renewable substrates. Molasses, a by-product of the sugar industry, contains mostly sucrose, a disaccharide composed of glucose and fructose, both easily absorbed by microorganisms. Yarrowia lipolytica, a platform for the production of various chemicals, can be engineered for sucrose utilization by heterologous invertase expression, yet the problem of preferential use of glucose over fructose remains, as fructose consumption begins only after glucose depletion what significantly extends the bioprocesses. We investigated the role of hexose transporters and hexokinase (native and fructophilic) in this preference. Analysis of growth profiles and kinetics of monosaccharide utilization has proven that the glucose preference in Y. lipolytica depends primarily on the affinity of native hexokinase for glucose. Interestingly, combined overexpression of either hexokinase with hexose transporters significantly accelerated citric acid biosynthesis and enhanced pentose phosphate pathway leading to secretion of polyols (31.5 g/L vs. no polyols in the control strain). So far, polyol biosynthesis was efficient in glycerol-containing media. Moreover, overexpression of fructophilic hexokinase in combination with hexose transporters not only shortened this process to 48 h (84 h for the medium with glycerol) but also allowed to obtain 23% more polyols (40 g/L) compared to the glycerol medium (32.5 g/L).

The N.C.Yeastract and CommunityYeastract databases to study gene and genomic transcription regulation in non-conventional yeasts

ABSTRACT Responding to the recent interest of the yeast research community in non-Saccharomyces cerevisiae species of biotechnological relevance, the N.C.Yeastract (http://yeastract-plus.org/ncyeastract/) was associated to YEASTRACT + (http://yeastract-plus.org/). The YEASTRACT + portal is a curated repository of known regulatory associations between transcription factors (TFs) and target genes in yeasts. N.C.Yeastract gathers all published regulatory associations and TF-binding sites for Komagataellaphaffii (formerly Pichia pastoris), the oleaginous yeast Yarrowia lipolytica, the lactose fermenting species Kluyveromyces lactis and Kluyveromyces marxianus, and the remarkably weak acid-tolerant food spoilage yeast Zygosaccharomyces bailii. The objective of this review paper is to advertise the update of the existing information since the release of N.C.Yeastract in 2019, and to raise awareness in the community about its potential to help the day-to-day work on these species, exploring all the information available in the global YEASTRACT + portal. Using simple and widely used examples, a guided exploitation is offered for several tools: (i) inference of orthologous genes; (ii) search for putative TF binding sites and (iii) inter-species comparison of transcription regulatory networks and prediction of TF-regulated networks based on documented regulatory associations available in YEASTRACT + for well-studied species. The usage potentialities of the new CommunityYeastract platform by the yeast community are also discussed.

Overexpression of YALI0B07117g Encoding Erythrose Reductase Homolog Results in Enhanced Erythritol Synthesis From Glycerol by the Yeast Yarrowia Lipolytica.

BackgroundPolyols are a group of sweet alcohols, frequently used as food additives. The constantly rising demand for polyols requires the application of new strategies to increase the production. Erythritol is synthesized by the yeast Yarrowia lipolytica under high osmotic pressure as an osmoprotectant. The metabolic pathway resulting in erythritol production remains partially unknown. However, the last reaction resulting in erythritol synthesis is conducted by an erythrose reductase (ER).ResultsThe Y. lipolytica strain was genetically modified to increase the erythritol yield and productivity, using glycerol as a sole carbon source. The modification focused on the ER homologue YALI0B07117g after the in silico analysis of the protein sequences of all reported ER homologues. Initial results in shake-flask experiments proved the influence of the gene YALI0B07117g in erythritol synthesis. Deletion of the gene resulted in 3-fold and 2-fold increased production of mannitol and arabitol, respectively. Overexpression of the native ER homologue gene showed a positive influence on erythritol production. Bath cultures were conducted and the obtained strain reached the yield of 0.4 g/g. The specific consumption rate (qs) increased from 5.83 g/g/L for the WT strain to 8.49 g/g/L for the engineered strain, while the productivity of erythritol increased from 0.28 g/L/h for the A101 strain to 0.41 g/L/h for the modified strain.ConclusionsOverexpression of the gene YALI0B07117g resulted in increased production of erythritol in the yeast Y. lipolytica. Disruption of the metabolic pathway by deletion of the gene results in higher production titers of mannitol and arabitol. Application of the research may prove positive for shortening the cultivation time due to the increased consumption rate of the substrate combined with increased parameters of erythritol synthesis.

Use of Non-Conventional Yeast Yarrowia lipolytica in Treatment or Upgradation of Hydrophobic Industry Wastes

The review aims to summarize the current knowledge on the possibility of using non-conventional yeast species Yarrowia lipolytica in the treatment and upgradation of industry wastes. Importantly Y. lipolytica yeast is argued as generally recognized as safe species, what indicates the high application potential of the reviewed technologies. Special emphasis in the paper was given on microbial processing of the food industry wastes, including fish and animals’ wastes utilization. Yarrowia-based processing of waste cooking oil or oil-bearing plants wastewaters, such as palm oil mill effluents or olive mill wastewater was reviewed. Recent advances in biosynthesis of valuable metabolites (e.g. lipases or microbial oil) with simultaneous wastes utilization by Y. lipolytica are additionally discussed. The broad implications of the present paper are a part of sustainable development policy. Graphic Abstract