Yeast Flocculation—Sedimentation and Flotation

Stewart, Graham G.

doi:10.3390/fermentation4020028

Cited by 43 publications

(36 citation statements)

References 115 publications

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“…The WT, ydr131cΔ, atg1Δ and ydr131cΔatg1Δ cells grown in YPD medium overnight and observed for floc formation visually and using microscope. The dependence of the floc formation on EDTA was tested by adding the 30mM of EDTA (chelating agent) to growth medium (S tewart 2018).…”

Section: Methodsmentioning

confidence: 99%

Deletion of Autophagy geneATG1and F-box motif encoding geneYDR131Ctogether leads to Synthetic Growth Defects and Flocculation behaviour inSaccharomyces cerevisiae

Shoket

Parvez

Sharma

et al. 2020

Preprint

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Running title: GeneticInteractions between YDR131C and ATG1 shows ubiquitination and 12 autophagy pathway cross talk governing growth fitness 13 Abstract: 20 F-box motif encoding YDR131C is functionally uncharacterized gene which forms the complex 21 with the SCF-E3 ligase. The F-box motif containing proteins are involved in substrate 22 recruitment for the ubiquitination and subsequent degradation through 26S proteasome. 23 Autophagy gene, ATG1 (ULK1in human) is a well conserved serine-threonine kinase, required 24 for vesicle formation and cytoplasm to vacuole targeting pathway. Atg1p forms the complex 25 with Atg13p and Atg17p during autophagy. The understanding of crosstalk between ubiquitin 26 and autophagy pathways is crucial for synthetic lethality screen and drug targeting. Here we have 27 conducted the study for genetic interaction between uncharacterized YDR131C and ATG1 gene 28 representing both specific and non-specific protein degradation pathways. The single and double 29 gene knockout strains of YDR131Cand ATG1 genes were constructed in the BY4741 genetic 30 background and analysed for growth fitness. The strains were also evaluated for cellular growth 31 response in presence of hydroxyurea (HU), methyl methane sulfonate (MMS), and hydrogen 32 peroxide (H 2 O 2 ) stress causing agents by spot assay. The ydr131cΔatg1Δ showed the synthetic 33 growth defect phenotype with floc formation in rich medium which showed floc disruption in 34 presence of EDTA. The ydr131cΔatg1Δ cells showed the sensitivity to stress agents HU, MMS, 35 and H 2 O 2 when compared with ydr131cΔ, atg1Δ, and WT cells.. Based on the observations, we 36 report that YDR131C and ATG1 functions in parallel pathways for growth fitness and cellular 37 growth response to stress agents. Interestingly this study also revealed the crosstalk between 38 ubiquitination and autophagy pathways. The defects in both the pathways could lead to synthetic 39 growth defects which may have implication for the precision medicine initiatives. 40 41 107 2019). Briefly Wild type (BY4741) and deletion derivatives (WT, ydr131cΔ, atg1Δ and 108 ydr131cΔatg1Δ) were grown to log phase (OD 600 0.8-1.0) and equal number of cell were serially 109 6 diluted. From each dilution a 3µl aliquot was spotted onto agar plates containing YPD, YPD + 110 stress causing agents such as a hydroxyurea (200mM), MMS (0.035%) and hydrogen peroxide 111 (4mM). The plates were incubated at 30°C for 2-3 days and imaged. 112 113 Fluorescence Microscopy for Cell Wall and Nuclear status 114To compare the cell wall status of WT, ydr131cΔ, atg1Δ, and ydr131cΔatg1Δ cells, Calcofluor 115 white staining assay described in (PRINGLE 1991; PREECHASUTH et al. 2015; SHARMA et al. 116 2019) was adopted. Briefly, WT and mutant strains were grown overnight at 30ºC and next day 117 transferred to fresh culture in 1:10 ratio. Cells were grown to log phase and collected by 118 centrifugation. Further, cells were suspended in 100µl of solution containing Calcofluor white 119 (50 μg/ml...

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Section: Methodsmentioning

confidence: 99%

Deletion of Autophagy geneATG1and F-box motif encoding geneYDR131Ctogether leads to Synthetic Growth Defects and Flocculation behaviour inSaccharomyces cerevisiae

Shoket

Parvez

Sharma

et al. 2020

Preprint

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“…The beneficial effect in oil recovery observed by the addition of CaCl 2 in centrifugation and not in GEOR can be explained by the fact that the calcium ions not only shield electrostatic repulsions but can also create links between the proteins surrounding the oil droplets. The creation of flocs of biomass when using CaCl 2 during fermentation broth has been demonstrated for Saccharomyces cerevisae (Nayyar et al, 2017;Stewart, 2018), however, no literature was found regarding E. coli fermentations with CaCl 2 flocculant. This seems to be confirmed by the microscopic pictures shown in Fig.…”

Section: Comparison Of Three Enhancement Oil Recovery Methods When Using Flocculants During Fermentation At Lab Scalementioning

confidence: 99%

Impact of flocculant addition in oil recovery from multiphasic fermentations

Basto

Jiménez

Mudde

et al. 2020

Food and Bioproducts Processing

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Emulsion formation is a major concern when dealing with multiphasic fermentations. Flocculants can be used together with other demulsification techniques to improve oil recovery in multiphasic fermentations. In this paper, the impact of adding flocculants during a multiphasic fermentation with 10 wt% dodecane, to destabilize the broth emulsion, improve creaming formation and enhance oil recovery is studied. Flocculants, CaCl 2 and (NH 4 ) 2 SO 4 were shown to be the most promising flocculants. Flocculant addition, their time of addition, and its impact on multiphasic fermentations has been evaluated by comparing fermentation performance against reference fermentations and three oil recovery methods: gravity settling, gas enhanced oil recovery and centrifugation. When adding 75 mM of (NH 4 ) 2 SO 4 during fermentation, the creaming rate during gravity settling increased 3-fold and the oil recovery by gas enhanced oil recovery was 35%, without altering fermentation performance.Addition of CaCl 2 during fermentation resulted in 88% and 67% oil recovery for early and late addition, which is a 4 and 3-fold increase in comparison with the reference. Yet, CaCl 2 deviated from standard fermentation performance when added immediately after second phase addition. In conclusion, flocculant addition during multiphasic fermentation can be used to destabilize microbial emulsions and potentially improve in situ oil recovery.

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“…involved in aging, oxidative stress response, and regulation of mitochondrial biogenesis [9] anchored to the cell by disulphide bridge [9] involved in remodeling of cell wall during culture development and stress/degradation responses [9] Sim1 cell wall remodeling during culture development [10,11] hpf1p haze protective mannoprotein [12] ecm33 required for cell wall integrity and assembly of the mannoprotein outer layer of the cell wall [6] cwp1 linked to β-1,3 and β-1,6-glucan through a phosphodiester bond [5] Does not require gPi anchor [5] cis3 component of outer cell wall layer for cell wall stability and optimal growth [10] Flocculation proteins flo1 flocculation inhibited by mannose residues [13] flo5 loss of gene flo5, requires propagation culture to be replaced [13] flo9…”

Section: Cell Wall Mannoproteins Uth1mentioning

confidence: 99%

“…Small flocculation protein [13] flo11 involved in filamentous, chain formation growth and flor forming then flocculation [13] Table 4. lc-Qtof-mS instrument parameters.…”

Section: Cell Wall Mannoproteins Uth1mentioning

confidence: 99%

Unfilterable Beer Haze Part II: Identifying Suspect Cell Wall Proteins

Huismann¹,

Gormley²,

Dzait³

et al. 2021

Journal of the American Society of Brewing Chemists

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The use of various diagnostic techniques has been previously utilized in the assessment of a commercially available India Pale Ale with cases of sporadically occurring unfilterable haze. The results from Part 1 suggested that β-glucans and proteins were the cause of the unfilterable haze and it was postulated that cell wall mannoproteins may also be a culprit of the unfilterable beer haze. In this follow-up study, proteins from high haze and low haze beer samples were precipitated and assessed using SDS-PAGE. Polyphenol interferences observed on the SDS-PAGE indicated that protein purification and targeted analysis was required. Proteins from high haze and low haze samples were fractionated and qualitatively identified via LC-MS. A library was built from FASTA sequences of targeted yeast proteins to qualitatively analyze the high haze and low haze samples. The protein fractionation was successful at purifying and isolating proteins from high and low haze samples. Two protein peaks were observed in the high haze sample, while one protein peak was observed in the low haze sample. The targeted LC/MS analysis discovered the presence of yeast cell wall mannoproteins and flocculation proteins, particularly Flo1 and Flo9. Understanding the source of these hazes can provide an opportunity for brewers to mitigate against their formation by adjusting brewing and yeast management practices.

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Yeast Flocculation—Sedimentation and Flotation

Cited by 43 publications

References 115 publications

Deletion of Autophagy geneATG1and F-box motif encoding geneYDR131Ctogether leads to Synthetic Growth Defects and Flocculation behaviour inSaccharomyces cerevisiae

Deletion of Autophagy geneATG1and F-box motif encoding geneYDR131Ctogether leads to Synthetic Growth Defects and Flocculation behaviour inSaccharomyces cerevisiae

Impact of flocculant addition in oil recovery from multiphasic fermentations

Unfilterable Beer Haze Part II: Identifying Suspect Cell Wall Proteins

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