Dataset Information

From mannan to bioethanol: cell surface co-display of ?-mannanase and ?-mannosidase on yeast Saccharomyces cerevisiae.

ABSTRACT: BACKGROUND:Mannans represent the largest hemicellulosic fraction in softwoods and also serve as carbohydrate stores in various plants. However, the utilization of mannans as sustainable resources has been less advanced in sustainable biofuel development. Based on a yeast cell surface-display technology that enables the immobilization of multiple enzymes on the yeast cell walls, we constructed a recombinant Saccharomyces cerevisiae strain that co-displays ?-mannanase and ?-mannosidase; this strain is expected to facilitate ethanol fermentation using mannan as a biomass source. RESULTS:Parental yeast S. cerevisiae assimilated mannose and glucose as monomeric sugars, producing ethanol from mannose. We constructed yeast strains that express tethered ?-mannanase and ?-mannosidase; co-display of the two enzymes on the cell surface was confirmed by immunofluorescence staining and enzyme activity assays. The constructed yeast cells successfully hydrolyzed 1,4-?-d-mannan and produced ethanol by assimilating the resulting mannose without external addition of enzymes. Furthermore, the constructed strain produced ethanol from 1,4-?-d-mannan continually during the third batch of repeated fermentation. Additionally, the constructed strain produced ethanol from ivory nut mannan; ethanol yield was improved by NaOH pretreatment of the substrate. CONCLUSIONS:We successfully displayed ?-mannanase and ?-mannosidase on the yeast cell surface. Our results clearly demonstrate the utility of the strain co-displaying ?-mannanase and ?-mannosidase for ethanol fermentation from mannan biomass. Thus, co-tethering ?-mannanase and ?-mannosidase on the yeast cell surface provides a powerful platform technology for yeast fermentation toward the production of bioethanol and other biochemicals from lignocellulosic materials containing mannan components.

SUBMITTER: Ishii J

PROVIDER: S-EPMC5009545 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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From mannan to bioethanol: cell surface co-display of β-mannanase and β-mannosidase on yeast Saccharomyces cerevisiae.

Ishii Jun J Okazaki Fumiyoshi F Djohan Apridah Cameliawati AC Hara Kiyotaka Y KY Asai-Nakashima Nanami N Teramura Hiroshi H Andriani Ade A Tominaga Masahiro M Wakai Satoshi S Kahar Prihardi P Yopi Prasetya Bambang B Ogino Chiaki C Kondo Akihiko A

Biotechnology for biofuels 20160902 1

<h4>Background</h4>Mannans represent the largest hemicellulosic fraction in softwoods and also serve as carbohydrate stores in various plants. However, the utilization of mannans as sustainable resources has been less advanced in sustainable biofuel development. Based on a yeast cell surface-display technology that enables the immobilization of multiple enzymes on the yeast cell walls, we constructed a recombinant Saccharomyces cerevisiae strain that co-displays β-mannanase and β-mannosidase; th ...[more]

PMID: 27594915

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Project description:UnlabelledBackgroundWhile the ethanol production from biomass by consolidated bioprocess (CBP) is considered to be the most ideal process, simultaneous saccharification and fermentation (SSF) is the most appropriate strategy in practice. In this study, one-pot bioethanol production, including cellulase production, saccharification of cellulose, and ethanol production, was investigated for the conversion of biomass to biofuel by co-culture of two different microorganisms such as a hyper cellulase producer, Acremonium cellulolyticus C-1 and an ethanol producer Saccharomyces cerevisiae. Furthermore, the operational conditions of the one-pot process were evaluated for maximizing ethanol concentration from cellulose in a single reactor.ResultsEthanol production from cellulose was carried out in one-pot bioethanol production process. A. cellulolyticus C-1 and S. cerevisiae were co-cultured in a single reactor. Cellulase producing-medium supplemented with 2.5 g/l of yeast extract was used for productions of both cellulase and ethanol. Cellulase production was achieved by A. cellulolyticus C-1 using Solka-Floc (SF) as a cellulase-inducing substrate. Subsequently, ethanol was produced with addition of both 10%(v/v) of S. cerevisiae inoculum and SF at the culture time of 60 h. Dissolved oxygen levels were adjusted at higher than 20% during cellulase producing phase and at lower than 10% during ethanol producing phase. Cellulase activity remained 8-12 FPU/ml throughout the one-pot process. When 50-300 g SF/l was used in 500 ml Erlenmeyer flask scale, the ethanol concentration and yield based on initial SF were as 8.7-46.3 g/l and 0.15-0.18 (g ethanol/g SF), respectively. In 3-l fermentor with 50-300 g SF/l, the ethanol concentration and yield were 9.5-35.1 g/l with their yields of 0.12-0.19 (g/g) respectively, demonstrating that the one-pot bioethanol production is a reproducible process in a scale-up bioconversion of cellulose to ethanol.ConclusionA. cellulolyticus cells produce cellulase using SF. Subsequently, the produced cellulase saccharifies the SF, and then liberated reducing sugars are converted to ethanol by S. cerevisiae. These reactions were carried out in the one-pot process with two different microorganisms in a single reactor, which does require neither an addition of extraneous cellulase nor any pretreatment of cellulose. Collectively, the one-pot bioethanol production process with two different microorganisms could be an alternative strategy for a practical bioethanol production using biomass.

Project description:In the search for renewable sources of energy, bioethanol stands out as a benchmark biofuel because its production is based on a proven technological platform. Bioethanol is produced mainly from the fermentation of carbohydrates derived from agricultural feedstocks by the yeast Saccharomyces cerevisiae. One of the most widely adopted strains is PE-2, a heterothallic diploid naturally adapted to the sugar cane fermentation process used in Brazil. Here we report the molecular genetic analysis of a PE-2 derived diploid (JAY270), and the complete genome sequence of a haploid derivative (JAY291). The JAY270 genome is highly heterozygous (~2 SNPs per kilobase), and has several structural polymorphisms between homologous chromosomes. These chromosomal rearrangements are confined to the peripheral regions of the chromosomes, and appear to reflect ectopic homologous recombination between repetitive DNA sequences. Despite the complex karyotype of JAY270, this diploid, when sporulated, had a high frequency of viable spores (~93%). Crosses of haploids derived from JAY270 to a haploid of the unrelated laboratory strain S288c also resulted in diploids that had good spore viability (75-95%). Thus, the rearrangements that exist near the ends of chromosomes do not impair meiosis and spore viability, as they do not span regions that contain essential genes. This observation is consistent with a model in which the peripheral regions of chromosomes represent plastic domains of the genome that are free to recombine ectopically and experiment with alternative structures that may be associated with a fitness benefit. We also explore features of the JAY270 and JAY291 genomes that help explain their high adaptation to industrial environments, exhibiting desirable phenotypes such as high cell mass production and fermentation kinetics, high temperature growth and oxidative stress tolerance. The genomic manipulation of such strains could enable the creation a new generation of industrial organisms, ideally suited for use as delivery vehicles for future bioenergy technologies.

Dataset Information

From mannan to bioethanol: cell surface co-display of ?-mannanase and ?-mannosidase on yeast Saccharomyces cerevisiae.

Publications

From mannan to bioethanol: cell surface co-display of β-mannanase and β-mannosidase on yeast Saccharomyces cerevisiae.

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