Lactic Acid Tolerance Research Articles

Characteristics and Functions of Dominant Yeasts Together with Their Applications during Strong-Flavor Baijiu Brewing.

Yeasts are pivotal brewing microbes that are associated with the flavor and quality of Chinese baijiu, yet research on dominant yeasts in strong-flavor baijiu brewing remains limited. In this study, Saccharomyces cerevisiae, Pichia kudriavzevii, and Kazachstania bulderi were identified as predominated yeasts in strong-flavor baijiu. Each strain showed distinct characteristics in ethanol resistance, thermal tolerance, and lactic acid tolerance, severally. S. cerevisiae FJ1-2 excelled in ethanol and ethyl ester production, P. kudriavzevii FJ1-1 in ethyl acetate, and K. bulderi FJ1-3 in lactic acid generation. Subsequently, the reinforced Fuqu of each yeast were severally prepared for application in baijiu brewing to verify their functions. Results revealed that the relative abundance of fortified yeast in each group rose. Pichia, Kazachstania, and Saccharomyces emerged as the core microbe for each group, respectively, by co-occurrence network analysis, influencing the microbiota to regulate flavor substances. In short, P. kudriavzevii FJ1-1 enhanced ethyl acetate. K. bulderi FJ1-3 improved ethyl caproate production and decreased levels of ethyl acetate and higher alcohols by modulating yeast community between Pichia and Saccharomyces. This is a systematic endeavor to study the functions of yeasts of strong-flavor baijiu, providing a solid basis for improving baijiu quality.

Evolution-aided improvement of the acid tolerance of Levilactobacillus brevis and its application in sourdough fermentation

Levilactobacillus brevis is crucial in food fermentation, particularly in sourdough production. However, the cultivation of L. brevis faces a challenge with accumulation of lactic acid, a major inhibitor. We aimed to increase the acid tolerance of L. brevis, an industrial strain for sourdough fermentation. We used the adaptive laboratory evolution (ALE) to obtain lactic acid tolerant strains. The evolved strain’s fermentation and metabolite profiles, alongside sensory evaluation, were compared with the parental strain by using various analytical techniques. The ALE approach increased lactic acid tolerance in the evolved strain showing an increased growth rate by 1.1 and 1.9 times higher than the parental strain at pH 4.1 and 6.5, respectively. Comprehensive analyses demonstrated its potential application in sourdough fermentation, promising reduced downstream costs. The evolved strain, free from genetically modified organisms concerns, has great potential for industrial use by exhibiting enhanced growth in acidic conditions without affecting consumers’ bread preferences.

Serine Improves Lactic Acid Stress Tolerance and Ethanol Production in Zygosaccharomyces bailii in Baijiu Fermentation.

Lactic acid is the primary inhibitor of the growth and ethanol production of yeasts in Baijiu fermentation. Certain amino acids have been found to be related to stress tolerance in yeasts. This study explored the effect of lactic acid stress on the ethanol-producing yeast Zygosaccharomyces bailii and evaluated the ability of serine to increase the lactic acid tolerance of Z. bailii in vitro. Serine significantly improved Z. bailii viability by 16.5% and ethanol production by 226.6% under lactic acid stress. Under lactic acid stress, serine supplementation led to an increase of 41.9% in cell wall integrity, 31.9% in cell membrane integrity, 296.6% in intracellular adenosine triphosphate (ATP), and 18.4% in the mitochondrial membrane potential. Finally, field emission scanning electron microscopy (FESEM) indicated that serine supplementation maintained the cell shape and reduced cell leakage. This study revealed a novel lactic acid tolerance mechanism of core functional yeasts during Jiang-flavor Baijiu fermentation.

Using transporter to enhance the acid tolerance of Bacillus coagulans DSM1

As the precursor of polylactic acid (PLA), optically pure l-lactic acid production is attracting increasing attention. The accumulation of lactic acid during fermentation inhibits strain growth. Therefore, it is necessary to improve the acid tolerance of lactic acid producers. In this study, comparative transcriptomic analysis was performed to investigate the effects of transporters on lactic acid tolerance of Bacillus coagulans DSM1, which is an l-lactic acid producer. The genes with more than two-fold up-regulation in transcriptional profile were further verified using real-time PCR. The transcriptional levels of RS06895, RS10595, RS10595, RS00500, RS00500, RS10635 and RS10635 were enhanced during lactic acid fermentation. Strain overexpressing RS10595 exhibited a retarded cell growth and low lactic acid production at pH 6.0, but an improved lactic acid production at pH 4.6. This study may facilitate the investigation of the acid tolerance mechanism in B. coagulans DSM1, as well as the construction of efficient lactic acid producers.

Inhibition of biofilm formation and virulence factors of cariogenic oral pathogen Streptococcus mutans by natural flavonoid phloretin

ABSTRACT Objectives To evaluate the effect and mechanism of action of the flavonoid phloretin on the growth and sucrose-dependent biofilm formation of Streptococcus mutans. Methods Minimum inhibitory concentration, viability, and biofilm susceptibility assays were conducted to assess antimicrobial and antibiofilm effect of phloretin. Biofilm composition and structure were analysed with scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). Water-soluble (WSG) and water-insoluble glucan (WIG) were determined using anthrone method. Lactic acid measurements and acid tolerance assay were performed to assess acidogenicity and aciduricity. Reverse transcription quantitative PCR (RT-qPCR) was used to measure the expression of virulence genes essential for surface attachment, biofilm formation, and quorum sensing. Results Phloretin inhibited S. mutans growth and viability in a dose-dependent manner. Furthermore, it reduced gtfB and gtfC gene expression, correlating with the reduction of extracellular polysaccharides (EPS)/bacteria and WIG/WSG ratio. Inhibition of comED and luxS gene expression, involved in stress tolerance, was associated with compromised acidogenicity and aciduricity of S. mutans. Conclusions Phloretin exhibits antibacterial properties against S. mutans, modulates acid production and tolerance, and reduces biofilm formation. Clinical significance Phloretin is a promising natural compound with pronounced inhibitory effect on key virulence factors of the cariogenic pathogen, S. mutans.

Expanded roles of lactate-sensing LldR in transcription regulation of the Escherichia coli K-12 genome: lactate utilisation and acid resistance.

LldR is a lactate-responsive transcription factor (TF) that transcriptionally regulates the lldPRD operon consisting of lactate permease and lactate dehydrogenase. The lldPRD operon facilitates the utilisation of lactic acid in bacteria. However, the role of LldR in whole genomic transcriptional regulation, and the mechanism involved in adaptation to lactate remains unclear. We used genomic SELEX (gSELEX) to comprehensively analyse the genomic regulatory network of LldR to understand the overall regulatory mechanism of lactic acid adaptation of the model intestinal bacterium Escherichia coli. In addition to the involvement of the lldPRD operon in utilising lactate as a carbon source, genes related to glutamate-dependent acid resistance and altering the composition of membrane lipids were identified as novel targets of LldR. A series of in vitro and in vivo regulatory analyses led to the identification of LldR as an activator of these genes. Furthermore, the results of lactic acid tolerance tests and co-culture experiments with lactic acid bacteria suggested that LldR plays a significant role in adapting to the acid stress induced by lactic acid. Therefore, we propose that LldR is an l-/d-lactate sensing TF for utilising lactate as a carbon source and for resistance to lactate-induced acid stress in intestinal bacteria.

Importance and mechanisms of S-adenosylmethionine and folate accumulation in sake yeast.

Sake yeasts have a range of brewing characteristics that are particularly beneficial for sake making including high ethanol fermentability, high proliferative capacity at low temperatures, lactic acid tolerance, and high ester productivity. On the other hand, sake yeasts also accumulate a diverse range of functional components. For example, significantly greater accumulation of S-adenosylmethionine (SAM), a compound that plays important regulatory roles in a range of biological processes as a major donor of methyl groups, occurs in sake yeasts compared to other microorganisms. Significantly greater accumulation of folate, a bioactive water-soluble vitamin (vitamin B9), also occurs in sake yeasts compared to laboratory yeasts, and the methyl group on SAM is supplied by folate. Accordingly, fully characterizing 'sake yeast identity' requires detailed understanding of the mechanisms underlying both the nutritional characteristics (functional components) and the brewing characteristics in sake yeasts. Therefore, this mini-review focuses on the accumulation of SAM and folate in sake yeast including descriptions of the genes known to contribute to SAM and folate accumulation and the underlying mechanisms.

Effect of dietary sodium bicarbonate on L.D.H enzyme level Female futsal players' lactic acid tolerance and exceptional endurance

The researcher has highlighted the importance of taking sodium bicarbonate for the level of LDH enzyme and lactic acid and the special endurance of futsal players, as sodium bicarbonate is one of the fluids that has a great effect on the athlete in terms of physical and physiological level, providing adequate physical fitness and increasing the activity and vitality of the body, helping the body to resist the nervous fatigue to which it is exposed. The study's goals included determining how consuming sodium bicarbonate affected the levels of lactic acid, the LDH enzyme, and the unique endurance of female futsal players (strength endurance, speed endurance, performance endurance). The researchers collected data using a variety of instruments, techniques, measurements, and qualifying methods while also using an experimental design with two experimental groups and a pre- and post-test. The statistical approach was adopted by the researcher (SPSS).

L-Lactic Acid Production Using Engineered Saccharomyces cerevisiae with Improved Organic Acid Tolerance.

Lactic acid is mainly used to produce bio-based, bio-degradable polylactic acid. For industrial production of lactic acid, engineered Saccharomyces cerevisiae can be used. To avoid cellular toxicity caused by lactic acid accumulation, pH-neutralizing agents are used, leading to increased production costs. In this study, lactic acid-producing S. cerevisiae BK01 was developed with improved lactic acid tolerance through adaptive laboratory evolution (ALE) on 8% lactic acid. The genetic basis of BK01 could not be determined, suggesting complex mechanisms associated with lactic acid tolerance. However, BK01 had distinctive metabolomic traits clearly separated from the parental strain, and lactic acid production was improved by 17% (from 102 g/L to 119 g/L). To the best of our knowledge, this is the highest lactic acid titer produced by engineered S. cerevisiae without the use of pH neutralizers. Moreover, cellulosic lactic acid production by BK01 was demonstrated using acetate-rich buckwheat husk hydrolysates. Particularly, BK01 revealed improved tolerance against acetic acid of the hydrolysates, a major fermentation inhibitor of lignocellulosic biomass. In short, ALE with a high concentration of lactic acid improved lactic acid production as well as acetic acid tolerance of BK01, suggesting a potential for economically viable cellulosic lactic acid production.

Mechanisms underlying lactic acid tolerance and its influence on lactic acid production in Saccharomyces cerevisiae.

One of the major bottlenecks in lactic acid production using microbial fermentation is the detrimental influence lactic acid accumulation poses on the lactic acid producing cells. The accumulation of lactic acid results in many negative effects on the cell such as intracellular acidification, anion accumulation, membrane perturbation, disturbed amino acid trafficking, increased turgor pressure, ATP depletion, ROS accumulation, metabolic dysregulation and metal chelation. In this review, the manner in which Saccharomyces cerevisiae deals with these issues will be discussed extensively not only for lactic acid as a singular stress factor but also in combination with other stresses. In addition, different methods to improve lactic acid tolerance in S. cerevisiae using targeted and non-targeted engineering methods will be discussed.

Improvement of lactic acid tolerance by cocktail δ-integration strategy and identification of the transcription factor PDR3 responsible for lactic acid tolerance in yeast Saccharomyces cerevisiae.

Although, yeast Saccharomyces cerevisiae is expected to be used as a host for lactic acid production, improvement of yeast lactic acid tolerance is required for efficient non-neutralizing fermentation. In this study, we optimized the expression levels of various transcription factors to improve the lactic acid tolerance of yeast by a previously developed cocktail δ-integration strategy. By optimizing the expression levels of various transcription factors, the maximum D-lactic acid production and yield under non-neutralizing conditions were improved by 1.2. and 1.6 times, respectively. Furthermore, overexpression of PDR3, which is known as a transcription factor involved in multi-drug resistance, effectively improved lactic acid tolerance in yeast. In addition, we clarified for the first time that high expression of PDR3 contributes to the improvement of lactic acid tolerance. PDR3 is considered to be an excellent target gene for studies on yeast stress tolerance and further researches are desired in the future.

Human Pathogenic Candida Species Respond Distinctively to Lactic Acid Stress

Several Candida species are opportunistic human fungal pathogens and thrive in various environmental niches in and on the human body. In this study we focus on the conditions of the vaginal tract, which is acidic, hypoxic, glucose-deprived, and contains lactic acid. We quantitatively analyze the lactic acid tolerance in glucose-rich and glucose-deprived environment of five Candida species: Candida albicans, Candida glabrata, Candida parapsilosis, Candida krusei and Candida tropicalis. To characterize the phenotypic space, we analyzed 40–100 clinical isolates of each species. Each Candida species had a very distinct response pattern to lactic acid stress and characteristic phenotypic variability. C. glabrata and C. parapsilosis were best to withstand high concentrations of lactic acid with glucose as carbon source. A glucose-deprived environment induced lactic acid stress tolerance in all species. With lactate as carbon source the growth rate of C. krusei is even higher compared to glucose, whereas the other species grow slower. C. krusei may use lactic acid as carbon source in the vaginal tract. Stress resistance variability was highest among C. parapsilosis strains. In conclusion, each Candida spp. is adapted differently to cope with lactic acid stress and resistant to physiological concentrations.

Construction of lactic acid-tolerant Saccharomyces cerevisiae by using CRISPR-Cas-mediated genome evolution for efficient D-lactic acid production.

Lactic acid (LA) is chemically synthesized or fermentatively produced using glucose as substrate, mainly using lactic acid bacteria. Polylactic acid is used as a biodegradable bioplastic for packaging materials, medical materials, and filaments for 3D printers. In this study, we aimed to construct a LA-tolerant yeast to reduce the neutralization cost in LA production. The pHLA2-51 strain was obtained through a previously developed genome evolution strategy, and transcriptome analysis revealed the gene expression profile of the mutant yeast. Furthermore, the expression of the genes associated with glycolysis and the LA synthesis pathway in the LA-tolerant yeast was comprehensively and randomly modified to construct a D-LA-producing, LA-tolerant yeast. In detail, DNA fragments expressing thirteen genes, HXT7, HXK2, PGI1, PFK1, PFK2, FBA1, TPI1, TDH3, PGK1, GPM1, ENO2, and PYK2, and D-lactate dehydrogenase (D-LDH) from Leuconostoc mesenteroides were randomly integrated into the genomic DNA in the LA-tolerant yeast. The resultant engineered yeast produced about 33.9 g/L of D-LA from 100 g/L glucose without neutralizing agents in a non-neutralized condition and 52.2 g/L of D-LA from 100 g/L glucose with 20 g/L CaCO3 in a semi-neutralized condition. Our research provides valuable insights into non-neutralized fermentative production of LA. KEY POINTS: • Lactic acid (LA) tolerance of yeast was improved by genome evolution. • The transcription levels of 751 genes were changed under LA stress. • Rapid LA production with semi-neutralization was achieved by modifying glycolysis. • A versatile yeast strain construction method based on the CRISPR system was proposed.

Sugar Metabolism of Scardovia wiggsiae, a Novel Caries-Associated Bacterium.

Scardovia wiggsiae has been detected from caries in children and adolescents and has been suggested to be a caries-associated microorganism. To investigate the cariogenic potential of S. wiggsiae, we examined carbohydrate metabolism and acid productivity, the fluoride sensitivity of carbohydrate metabolism and the mechanism by which fluoride inhibits carbohydrate metabolism, and the acid sensitivity of carbohydrate metabolism in this bacterium. S. wiggsiae metabolized glucose and reduced the environmental pH to 3.5. It mainly produced acetic acid from glucose, together with small amounts of lactic and formic acid. The 50% inhibitory concentration of fluoride for acid production was 8.0 mM at pH 7.0 and 1.5 mM at pH 5.5, which were much higher than those of representative caries-associated bacteria, such as Streptococcus mutans. Metabolomic profiles showed the accumulation of 3-phosphoglycerate and a marked reduction in the pyruvate concentration in the presence of fluoride, suggesting that fluoride inhibits the latter half of glycolysis, including enolase activity. Enolase activity was inhibited by fluoride in S. wiggsiae, but it was more fluoride-tolerant than the enolase activity of S. mutans. Unlike in S. mutans, lactic acid did not inhibit acid production by S. wiggsiae at acidic pH. These results indicate that S. wiggsiae exhibits high acid production and tolerance to fluoride and lactic acid. S. wiggsiae possesses a unique metabolic pathway, the F6PPK shunt, which might allow it to avoid the lactate-formate pathway, including fluoride-sensitive enolase activity, and enable metabolic flow to the fluoride-tolerant acetate pathway. The fluoride tolerance of S. wiggsiae’s enolase activity also increases the fluoride tolerance of its carbohydrate metabolism. The lactic acid tolerance of S. wiggsiae’s acid production might result in S. wiggsiae having high acidogenic and aciduric potential and make it ecologically competitive in acidic environments, such as caries lesions, where lactic acid predominates.

Cooperative Response of Pichia kudriavzevii and Saccharomyces cerevisiae to Lactic Acid Stress in Baijiu Fermentation.

Lactic acid is a universal metabolite, as well as a growth inhibitor of ethanol producers in Baijiu fermentation. Revealing the mechanism of lactic acid tolerance is essential for the yield of fermented foods. Here, we employed reverse transcription-quantitative polymerase chain reaction to explore the degradation mechanism of lactic acid, based on the coculture of Pichia kudriavzevii and Saccharomyces cerevisiae. Under high lactic acid stress, P. kudriavzevii decreased lactic acid from 40.00 to 35.46 g L-1 within 24 h. Then, S. cerevisiae restored its capacity to degrade lactic acid. Finally, lactic acid decreased to 26.29 g L-1. Coculture significantly enhanced lactic acid consumption compared to the monoculture of P. kudriavzevii (90% higher) or S. cerevisiae (209% higher). We found that lactate catabolism, H+ extrusion, and glycerol transport were the lactic acid tolerance pathways in yeasts. This study reveals the novel acid tolerance mechanisms of microbiota and would provide new strategies for ethanol production under acid stress.

Low-pH production of d-lactic acid using newly isolated acid tolerant yeast Pichia kudriavzevii NG7.

Lactic acid is a platform chemical for the sustainable production of various materials. To develop a robust yeast platform for low-pH production of d-lactic acid (LA), an acid-tolerant yeast strain was isolated from grape skins and named Pichia kudriavzevii NG7 by ribosomal RNA sequencing. This strain could grow at pH 2.0 and 50°C. For the commercial application of P. kudriavzevii NG7 as a lactic acid producer, the ethanol fermentation pathway was redirected to lactic acid by replacing the pyruvate decarboxylase 1 gene (PDC1) with the d-lactate dehydrogenase gene (d-LDH) derived from Lactobacillus plantarum. To enhance lactic acid tolerance, this engineered strain was adapted to high lactic acid concentrations, and a new transcriptional regulator, PAR1, responsible for acid tolerance, was identified by whole-genome resequencing. The final engineered strain produced 135 g/L and 154 g/L of d-LA with productivity over 3.66 g/L/hr at pH 3.6 and 4.16 g/L/hr at pH 4.7, respectively.

Transcriptional Response to Lactic Acid Stress in the Hybrid Yeast Zygosaccharomyces parabailii.

ABSTRACTLactic acid has a wide range of applications starting from its undissociated form, and its production using cell factories requires stress-tolerant microbial hosts. The interspecies hybrid yeast Zygosaccharomyces parabailii has great potential to be exploited as a novel host for lactic acid production, due to high organic acid tolerance at low pH and a fermentative metabolism with a high growth rate. Here we used mRNA sequencing (RNA-seq) to analyze Z. parabailii's transcriptional response to lactic acid added exogenously, and we explore the biological mechanisms involved in tolerance. Z. parabailii contains two homeologous copies of most genes. Under lactic acid stress, the two genes in each homeolog pair tend to diverge in expression to a significantly greater extent than under control conditions, indicating that stress tolerance is facilitated by interactions between the two gene sets in the hybrid. Lactic acid induces downregulation of genes related to cell wall and plasma membrane functions, possibly altering the rate of diffusion of lactic acid into cells. Genes related to iron transport and redox processes were upregulated, suggesting an important role for respiratory functions and oxidative stress defense. We found differences in the expression profiles of genes putatively regulated by Haa1 and Aft1/Aft2, previously described as lactic acid responsive in Saccharomyces cerevisiae. Furthermore, formate dehydrogenase (FDH) genes form a lactic acid-responsive gene family that has been specifically amplified in Z. parabailii in comparison to other closely related species. Our study provides a useful starting point for the engineering of Z. parabailii as a host for lactic acid production.IMPORTANCE Hybrid yeasts are important in biotechnology because of their tolerance to harsh industrial conditions. The molecular mechanisms of tolerance can be studied by analyzing differential gene expression under conditions of interest and relating gene expression patterns to protein functions. However, hybrid organisms present a challenge to the standard use of mRNA sequencing (RNA-seq) to study transcriptional responses to stress, because their genomes contain two similar copies of almost every gene. Here we used stringent mapping methods and a high-quality genome sequence to study the transcriptional response to lactic acid stress in Zygosaccharomyces parabailii ATCC 60483, a natural interspecies hybrid yeast that contains two complete subgenomes that are approximately 7% divergent in sequence. Beyond the insights we gained into lactic acid tolerance in this study, the methods we developed will be broadly applicable to other yeast hybrid strains.

PM 7/128 (1) Xanthomonas axonopodis pv. allii

Specific scopeThis Standard describes a diagnostic protocol for Xanthomonas axonopodis pv. allii.This Standard should be used in conjunction with PM 7/76 Use of EPPO diagnostic protocols.Specific approval and amendmentFirst approved in 2016‐09.

Long-term adaptive evolution of Leuconostoc mesenteroides for enhancement of lactic acid tolerance and production.

BackgroundLactic acid has been approved by the United States Food and Drug Administration as Generally Regarded As Safe (GRAS) and is commonly used in the cosmetics, pharmaceutical, and food industries. Applications of lactic acid have also emerged in the plastics industry. Lactic acid bacteria (LAB), such as Leuconostoc and Lactobacillus, are widely used as lactic acid producers for food-related and biotechnological applications. Nonetheless, industrial mass production of lactic acid in LAB is a challenge mainly because of growth inhibition caused by the end product, lactic acid. Thus, it is important to improve acid tolerance of LAB to achieve balanced cell growth and a high titer of lactic acid. Recently, adaptive evolution has been employed as one of the strategies to improve the fitness and to induce adaptive changes in bacteria under specific growth conditions, such as acid stress.ResultsWild-type Leuconostoc mesenteroides was challenged long term with exogenously supplied lactic acid, whose concentration was increased stepwise (for enhancement of lactic acid tolerance) during 1 year. In the course of the adaptive evolution at 70 g/L lactic acid, three mutants (LMS50, LMS60, and LMS70) showing high specific growth rates and lactic acid production were isolated and characterized. Mutant LMS70, isolated at 70 g/L lactic acid, increased d-lactic acid production up to 76.8 g/L, which was twice that in the wild type (37.8 g/L). Proteomic, genomic, and physiological analyses revealed that several possible factors affected acid tolerance, among which a mutation of ATPase ε subunit (involved in the regulation of intracellular pH) and upregulation of intracellular ammonia, as a buffering system, were confirmed to contribute to the observed enhancement of tolerance and production of d-lactic acid.ConclusionsDuring adaptive evolution under lethal stress conditions, the fitness of L. mesenteroides gradually increased to accumulate beneficial mutations according to the stress level. The enhancement of acid tolerance in the mutants contributed to increased production of d-lactic acid. The observed genetic and physiological changes may systemically help remove protons and retain viability at high lactic acid concentrations.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0662-3) contains supplementary material, which is available to authorized users.

GSF2 deletion increases lactic acid production by alleviating glucose repression in Saccharomyces cerevisiae.

Improving lactic acid (LA) tolerance is important for cost-effective microbial production of LA under acidic fermentation conditions. Previously, we generated LA-tolerant D-LA-producing S. cerevisiae strain JHY5310 by laboratory adaptive evolution of JHY5210. In this study, we performed whole genome sequencing of JHY5310, identifying four loss-of-function mutations in GSF2, SYN8, STM1, and SIF2 genes, which are responsible for the LA tolerance of JHY5310. Among the mutations, a nonsense mutation in GSF2 was identified as the major contributor to the improved LA tolerance and LA production in JHY5310. Deletion of GSF2 in the parental strain JHY5210 significantly improved glucose uptake and D-LA production levels, while derepressing glucose-repressed genes including genes involved in the respiratory pathway. Therefore, more efficient generation of ATP and NAD+ via respiration might rescue the growth defects of the LA-producing strain, where ATP depletion through extensive export of lactate and proton is one of major reasons for the impaired growth. Accordingly, alleviation of glucose repression by deleting MIG1 or HXK2 in JHY5210 also improved D-LA production. GSF2 deletion could be applied to various bioprocesses where increasing biomass yield or respiratory flux is desirable.