Yeast Protein Research Articles

Prostate secretory protein 94 inhibits sterol binding and export by the mammalian CAP protein CRISP2 in a calcium-sensitive manner

Members of the CAP protein superfamily are present in all kingdoms of life and have been implicated in many different processes, including pathogen defense, immune evasion, sperm maturation, and cancer progression. Most CAP proteins are secreted glycoproteins and share a unique conserved αβα sandwich fold. The precise mode of action of this class of proteins, however, has remained elusive. Saccharomyces cerevisiae has three CAP family members, termed pathogen related in yeast (Pry). We have previously shown that Pry1 and Pry2 export sterols in vivo and that they bind sterols in vitro. This sterol binding and export function of yeast Pry proteins is conserved in the mammalian CRISP proteins and other CAP superfamily members. CRISP3 is an abundant protein of the human seminal plasma and interacts with prostate secretory protein of 94 amino acids (PSP94), another major protein component in the seminal plasma. Here we examine whether the interaction between CRISP proteins and PSP94 affects the sterol binding function of CAP family members. We show that coexpression of PSP94 with CAP proteins in yeast abolished their sterol export function and the interaction between PSP94 and CAP proteins inhibits sterol binding in vitro. In addition, mutations that affect the formation of the PSP94–CRISP2 heteromeric complex restore sterol binding. Of interest, we found the interaction of PSP94 with CRISP2 is sensitive to high calcium concentrations. The observation that PSP94 modulates the sterol binding function of CRISP2 in a calcium-dependent manner has potential implications for the role of PSP94 and CRISP2 in prostate physiology and progression of prostate cancer.

Yeast Protein as an Easily Accessible Food Source.

In recent years, the awareness and willingness of consumers to consume healthy food has grown significantly. In order to meet these needs, scientists are looking for innovative methods of food production, which is a source of easily digestible protein with a balanced amino acid composition. Yeast protein biomass (single cell protein, SCP) is a bioavailable product which is obtained when primarily using as a culture medium inexpensive various waste substrates including agricultural and industrial wastes. With the growing population, yeast protein seems to be an attractive alternative to traditional protein sources such as plants and meat. Moreover, yeast protein biomass also contains trace minerals and vitamins including B-group. Thus, using yeast in the production of protein provides both valuable nutrients and enhances purification of wastes. In conclusion, nutritional yeast protein biomass may be the best option for human and animal nutrition with a low environmental footprint. The rapidly evolving SCP production technology and discoveries from the world of biotechnology can make a huge difference in the future for the key improvement of hunger problems and the possibility of improving world food security. On the market of growing demand for cheap and environmentally clean SCP protein with practically unlimited scale of production, it may soon become one of the ingredients of our food. The review article presents the possibilities of protein production by yeast groups with the use of various substrates as well as the safety of yeast protein used as food.

Cyberlindnera jadinii yeast as a protein source in early- to mid-lactation dairy cow diets: Effects on feed intake, ruminal fermentation, and milk production

We examined the effects of substituting soybean meal with either yeast protein from Cyberlindnera jadinii or barley in concentrate feeds on feed intake, ruminal fermentation products, milk production, and milk composition in Norwegian Red (NRF) dairy cows. The concentrate feeds were prepared in pellet form as soy-based (SBM; where soybean meal is included as a protein ingredient), yeast-based (YEA; soybean meal replaced with yeast protein), or barley-based (BAR; soybean meal replaced with barley). The SBM contained 7.0% soybean meal on a dry matter (DM) basis. This was replaced with yeast protein and barley in the YEA and BAR concentrate feeds, respectively. A total of 48 early- to mid-lactation [days in milk ± standard deviation (SD): 103 ± 33.5 d] NRF cows in their first to fourth parity and with initial milk yield of 32.6 kg (SD = 7.7) were allocated into 3 groups, using a randomized block design, after feeding a common diet [SBM and good-quality grass silage: crude protein (CP) and neutral detergent fiber (NDF) content of 181 and 532 g/kg of DM, respectively] for 14 d (i.e., covariate period). The groups (n = 16) were then fed one of the dietary treatments (SBM, YEA, or BAR) for a period of 56 d (i.e., experimental period). The concentrate feeds were offered in split portions from 3 automatic feeders using electronic identification, with ad libitum access to the same grass silage. Dietary treatments had no effect on daily silage intake, total DM intake, or total NDF intake. Dietary CP intake was lower and starch intake was higher in the BAR group compared with the other groups. Ruminal fluid pH, short-chain volatile fatty acid (VFA) concentrations, acetate-to-propionate ratio, and non-glucogenic to glucogenic VFA ratio were not affected by dietary treatments. No effects of the dietary treatments were observed on body weight change, body condition score change, milk yield, energy-corrected milk yield, milk lactose and fat percentages, or their yields. In conclusion, yeast protein can substitute conventional soybean meal in dairy cow diets without adverse effect on milk production and milk composition, given free access to good-quality grass silage.

Corn distillers solubles as a novel bioresource of bioactive peptides with ACE and DPP IV inhibition activity: characterization, in silico evaluation, and molecular docking.

This study aimed to investigate the biological potential of underutilized and low-value corn distillers solubles, containing a unique unexplored blend of heat-treated corn and yeast proteins, from the bioethanol industries, by bioinformatic and biochemical approaches. Protein hydrolysates were produced by applying four commercially accessible proteases, among which alcalase provided the best results in terms of yield, degree of hydrolysis, molecular weight, number of proteins, bioactive peptides, and deactivation against anti-angiotensin I-converting enzyme (ACE) and anti-dipeptidyl peptidase IV (DPP IV). The optimal conditions to produce anti-ACE and anti-DPP IV peptides were using alcalase for 10.82 h and an enzyme : substrate ratio of 7.90 (%w/w), with inhibition values for ACE and DPP IV of 98.76 ± 1.28% and 34.99 ± 1.44%, respectively. Corn (α-zein) and yeast (glyceraldehyde-3-phosphate dehydrogenase) proteins were mainly suitable, upon enzymolysis, for the release of bioactive peptides. The peptides DPANLPWG, FDFFDNIN, WNGPPGVF, and TPPFHLPPP inhibited ACE more effectively as verified with binding energies of -11.3, -11.6, -10.5, and -11.6 kcal mol-1, respectively, as compared to captopril (-6.38 kcal mol-1). Compared with the binding energy of sitagliptin (-8.6 kcal mol-1), WNGPPGVF (-9.6 kcal mol-1), WPLPPFG (-9.8 kcal mol-1), LPPYLPS (-9.7 kcal mol-1), TPPFHLPPP (-10.1 kcal mol-1), and DPANLPWG peptides (-10.1 kcal mol-1) had greater inhibition potential against DPP IV. The peptides impeded ACE and DPP IV majorly via hydrophobic and hydrogen linkage interactions. The key amino acids TYR523, GLU384, and HIS353 were bound to the catalytic sites of ACE and GLN553, GLU206, PHE364, VAL303, and THR304 were bound to the DPP IV enzyme. The PHs can be used as ingredients in the feed or food industries with possible health advantages.

An Aptamer-based mRNA Affinity Purification Procedure (RaPID) for the Identification of Associated RNAs (RaPID-seq) and Proteins (RaPID-MS) in Yeast.

RNA-RNA and RNA-protein interactions are involved in the regulation of gene expression. Here, we describe an updated and extended version of our RNA purification and protein identification (RaPID) protocol for the pulldown of aptamer-tagged mRNAs by affinity purification. The method takes advantage of the high affinity interaction between the MS2 RNA aptamer and the MS2 coat protein (MCP), as well as that between streptavidin-binding peptide (SBP) and streptavidin. Thus, it employs MCP-SBP fusions to affinity purify MS2-tagged target RNAs of interest over immobilized streptavidin. Purified aptamer-tagged mRNAs, along with any associated RNAs and proteins, are then sent for RNA sequencing (RaPID-seq) or mass spectrometry (RaPID-MS), which allows for the identification of bound cohort RNAs and proteins, respectively.

Optimization of Yeast Cultivation Factors for Improved SCP Production

Abstract Yeast Single-Cell Proteins (SCP) production using various agro-industrial byproducts and wastes have significant potential as an alternative to the soy meal, and fish meal protein used for livestock and aquaculture feeds. The use of organic wastes as a substrate in the fermentation processes can be accepted as one of the solutions to reduce the total price of the culture and an environmentally friendlier method of removing these residues. This review article focuses on the yeast biomass yield and protein content increase strategies, which is impossible without understanding metabolic pathways and switching mechanisms. The present work discusses optimization strategies for protein-enriched yeast biomass production, such as fermentation medium composition, including a selection of carbon and nitrogen sources and their ratio, supplemented trace elements, and cultivation conditions such as pH, temperature, time of cultivation, and inoculum size. This review summarizes the theoretical knowledge and experimental results of other researchers that provide an overview of the achievements of the last decades in the production of SCP.

Cassava pulp can be nutritionally improved by yeast and various crude protein levels fed to cattle

Context Dietary supplementation especially feed residues improve by yeast affected rumen fermentation. Aims The aim of the present experiment was to determine the nutritive value, fermentation efficiency and rumen ecology of yeast-fermented cassava pulp, under the use of two levels of protein in concentrate mixtures, in beef cattle. Methods Four beef cattle, 2–3 years of age, were randomly assigned to the following treatments according to a 2 × 2 factorial arrangement in a 4 × 4 Latin square design: cassava pulp fermented either with baker’s yeast or LDD 6 (Factor A), with 16% or 18% CP in concentrate (Factor B). Key results CP digestibility and blood urea nitrogen were higher in animals fed both baker’s yeast-fermented and LDD 6-fermented cassava pulp with 18% CP than in those fed baker’s yeast-fermented cassava pulp with 16% CP and LDD 6-fermented cassava pulp with 16% CP. The highest total volatile fatty acid (TVFA) and butyrate (C4) concentrations were in animals fed baker’s yeast-fermented cassava pulp with 16% CP. Moreover, CH4 production was at its lowest when animals were fed with baker’s yeast-fermented cassava pulp together with a concentrate with 16% CP. Animal’s fed baker’s yeast-fermented and LDD 6-fermented cassava pulp with 18% CP had higher concentrations of purine derivatives, higher microbial nitrogen supply and efficiency of microbial protein synthesis than did those fed either baker’s yeast-fermented or LDD 6-fermented cassava pulp with 16% CP. In addition, protozoa population in animals fed baker’s yeast-fermented cassava pulp was lower than in those fed LDD 6-fermented cassava pulp, and baker’s yeast-fermented cassava pulp with 16% CP had the lowest protozoa population. However, F. succinogenes was the highest in animals fed yeast-fermented and LDD 6-fermented cassava pulp with 18% CP. Conclusions It is concluded that yeast-fermented cassava pulp can be nutritionally improved, especially when used in combination with a concentrate of 16% CP to achieve the highest level of TVFA and C3 and to reduce rumen protozoal population and CH4 production. Implications The present study suggests that feeding animals with yeast-fermented cassava pulp with a concentrate of 16% CP could improve feed quality and, hence, beef animal performance in Thailand.

Protein complex prediction in large protein–protein interaction network

Due to high computational complexity, the detection of protein complexes in large protein–protein interaction (PPI) networks remains a challenging problem. Finding the actual protein complexes from a large PPI network requires a sophisticated algorithm. The protein complexes exhibit in densely connected sub-graphs in a PPI network. This paper presents a novel algorithm based on a metaheuristic method for protein complex prediction in large PPI networks. The algorithm mimics the density-based graph clustering method with biological heuristics to identify the protein complexes. The algorithm is enhanced by a local search algorithm and three repair operators. A new function has been developed for computing cluster density. The method was applied to the yeast and human protein interaction data and compared with the state-of-the-art algorithms. The comparisons demonstrate the best performance of the proposed algorithm in terms of accuracy and f-measure.

The Interaction of Yeast Protein with Hydrophilic and Hydrophobic Bioactive Molecules, the Binding, Structure, and Function Evaluation

The Interaction of Yeast Protein with Hydrophilic and Hydrophobic Bioactive Molecules, the Binding, Structure, and Function Evaluation

Serial fractionation of spent brewer's yeast protein hydrolysate by ultrafiltration: A peptide-rich product with low RNA content

Spent brewer's yeast (SBY) application as a new source of peptides depends on the development of proper downstream processes. This work aimed to separate yeast hydrolysate peptides from sugars and ribonucleic acids (RNA) by a 3-step designed fractionation process using ceramic ultra and nanofiltration following two sequences: (1) 50, 8 and 1 kDa and (2) 15, 8 and 1 kDa molecular weight cut-off (MWCO) membranes. In step one, 15 kDa MWCO membrane retained more components and increased performance of subsequent steps, when compared to 50 kDa. Peptide composition of the initial hydrolysate was gradually changed until 90% of the peptides smaller than 1 kg mol −1 were found in the final permeate. Fractionation increased peptide purity regarding RNA and total sugars up to 1.7 and 2.7-fold, respectively. Innovative protein-rich ingredients from SBY residue with an RNA content as low as 1.4% were produced by membrane separation technology. High quality ingredients presented different peptide profiles and great potential in food and pharmaceutical industries applications. • Efficient RNA reduction was achieved using membrane separation technology. • RNA content in protein-rich fraction was as low as 1.4% (dry weight). • Membranes of 15–1 kDa range were adapted to spent yeast ultrafiltration. • Protein purity of obtained fractions increased 1.7 and 2.7-fold for RNA and sugars. • A fraction with bioactive potential (65% of the peptides < 1 kg mol −1 ) was produced.

The Conserved Yeast Protein Knr4 Involved in Cell Wall Integrity is a Multi-Domain Intrinsically Disordered Protein

The Conserved Yeast Protein Knr4 Involved in Cell Wall Integrity is a Multi-Domain Intrinsically Disordered Protein

Secretome-based screening of fusion partners and their application in recombinant protein secretion in Saccharomyces cerevisiae.

For the efficient production of heterologous proteins in the yeast Saccharomyces cerevisiae, we screened for a novel fusion partner from the yeast secretome. From twenty major proteins identified from the yeast secretome, we selected Scw4p, a cell wall protein with similarity to glucanase, and modified to develop a general fusion partner for the secretory expression of heterologous proteins in yeast. The optimal size of the SCW4 gene to act as an efficient fusion partner was determined by C-terminal truncation analysis; two of the variants, S1 (truncated at codon 115Q) and S2 (truncated at codon 142E), were further used for the secretion of heterologous proteins. When fused with S2, the secretion of three target proteins (hGH, exendin-4, and hPTH) significantly increased. Conserved O-glycosylation sites (Ser/Thr-rich domain) and hydrophilic sequences of S2 were deemed important for the function of S2 as a secretion fusion partner. Approximately 5g/L of the S2-exendin-4 fusion protein was obtained from fed-batch fermentation. Intact target proteins were easily purified by affinity chromatography after in vitro processing of the fusion partner. This system may be of general application for the secretory production of heterologous proteins in S. cerevisiae. KEY POINTS : • Target proteins were efficiently secreted with their N-terminus fused to Scw4p. • O-glycosylation and hydrophilic stretches in Scw4p were important for protein secretion. • A variant of Scw4p (S2) was successfully applied for the secretory expression of heterologous proteins.

Identification of uncharacterized proteins potentially localized to mitochondria (UPMs) in Saccharomyces cerevisiae using a fluorescent protein unstable in the cytoplasm.

Eukaryotic cells are composed of organelles, and each organelle contains proteins that play a role in its function. Therefore, the localization of a protein, especially to organelles, is a clue to infer the function of that protein. In this study, we attempted to identify novel mitochondrially localized proteins in the budding yeast Saccharomyces cerevisiae using a fluorescent protein (GFPdeg) that is rapidly degraded in the cytoplasm. Of the budding yeast proteins predicted to localize to mitochondria by the prediction tool Deeploc-1.0, those with known mitochondrial localization or functional relevance were eliminated, and 95 proteins of unknown function were selected as candidates for analysis. By forced expression of GFPdeg fusion proteins with these proteins and observation of their localization, we identified 35 uncharacterized proteins potentially localized to mitochondria (UPMs) including 8 previously identified proteins that localize to mitochondria. Most of these had no N-terminal mitochondrial localization signal and were evolutionarily young "emerging genes" that exist only in S. cerevisiae. Some of these genes were found to be upregulated during the postdiauxic shift phase when mitochondria are being developed, suggesting that they are actually involved in some mitochondrial function.

Frameshifting at collided ribosomes is modulated by elongation factor eEF3 and by integrated stress response regulators Gcn1 and Gcn20.

Ribosome stalls can result in ribosome collisions that elicit quality control responses, one function of which is to prevent ribosome frameshifting, an activity that entails the interaction of the conserved yeast protein Mbf1 with uS3 on colliding ribosomes. However, the full spectrum of factors that mediate frameshifting during ribosome collisions is unknown. To delineate such factors in the yeast Saccharomyces cerevisiae, we used genetic selections for mutants that affect frameshifting from a known ribosome stall site, CGA codon repeats. We show that the general translation elongation factor eEF3 and the integrated stress response (ISR) pathway components Gcn1 and Gcn20 modulate frameshifting in opposing manners. We found a mutant form of eEF3 that specifically suppressed frameshifting, but not translation inhibition by CGA codons. Thus, we infer that frameshifting at collided ribosomes requires eEF3, which facilitates tRNA–mRNA translocation and E-site tRNA release in yeast and other single cell organisms. In contrast, we found that removal of either Gcn1 or Gcn20, which bind collided ribosomes with Mbf1, increased frameshifting. Thus, we conclude that frameshifting is suppressed by Gcn1 and Gcn20, although these effects are not mediated primarily through activation of the ISR. Furthermore, we examined the relationship between eEF3-mediated frameshifting and other quality control mechanisms, finding that Mbf1 requires either Hel2 or Gcn1 to suppress frameshifting with wild-type eEF3. Thus, these results provide evidence of a direct link between translation elongation and frameshifting at collided ribosomes, as well as evidence that frameshifting is constrained by quality control mechanisms that act on collided ribosomes.

Production of galactosylated complex-type N-glycans in glycoengineered Saccharomyces cerevisiae

N-glycosylation is an important posttranslational modification affecting the properties and quality of therapeutic proteins. Glycoengineering in yeast aims to produce proteins carrying human-compatible glycosylation, enabling the production of therapeutic proteins in yeasts. In this work, we demonstrate further development and characterization of a glycoengineering strategy in a Saccharomyces cerevisiae Δalg3 Δalg11 strain where a truncated Man3GlcNAc2 glycan precursor is formed due to a disrupted lipid-linked oligosaccharide synthesis pathway. We produced galactosylated complex-type and hybrid-like N-glycans by expressing a human galactosyltransferase fusion protein both with and without a UDP-glucose 4-epimerase domain from Schizosaccharomyces pombe. Our results showed that the presence of the UDP-glucose 4-epimerase domain was beneficial for the production of digalactosylated complex-type glycans also when extracellular galactose was supplied, suggesting that the positive impact of the UDP-glucose 4-epimerase domain on the galactosylation process can be linked to other processes than its catalytic activity. Moreover, optimization of the expression of human GlcNAc transferases I and II and supplementation of glucosamine in the growth medium increased the formation of galactosylated complex-type glycans. Additionally, we provide further characterization of the interfering mannosylation taking place in the glycoengineered yeast strain.Key points• Glycoengineered Saccharomyces cerevisiae can form galactosylated N-glycans.• Genetic constructs impact the activities of the expressed glycosyltransferases.• Growth medium supplementation increases formation of target N-glycan structure.

Recycling and Conversion of Yeasts into Organic Nitrogen Sources for Wine Fermentation: Effects on Molecular and Sensory Attributes

Organic nitrogen plays a significant role in the fermentation performance and production of esters and higher alcohols. This study assessed the use of yeast protein hydrolysate (YPH) as a nitrogen source for grape must fermentation. In this study, we prepared an enzymatic protein hydrolysate using yeasts recovered from a previous fermentation of wine. Three treatments were performed. DAP supplementation was used as a control, while two YPH treatments were used. Low (LDH) and high degrees of hydrolysis (HDH), 3.5% and 10%, respectively, were chosen. Gas chromatography and principal component analysis indicated a significant positive influence of YPH-supplementations on the production of esters and higher alcohols. Significantly high concentrations of 3-methyl-1-penthanol, isoamyl alcohol, isobutanol, and 2-phenylethanol were observed. Significant odorant activity was obtained for 3-methyl-1-pentanol and ethyl-2-hexenoate. The use of YPH as nitrogen supplementation is justified as a recycling yeasts technique by the increase in volatile compounds.

Isolation of novel wheat bran antifreeze polysaccharides and the cryoprotective effect on frozen dough quality

Different wheat bran antifreeze polysaccharides (WBAP) were first isolated by ice shell separation method. The composition and antifreeze activity of varied WBAP were characterized and their cryoprotective effects on frozen dough were comprehensively studied with the emphasis on ice crystals, yeast and gluten proteins. WBAP with enriched arabinoxylan (AX) of high arabinofuranose/xylopyranose ratio (A/X) showed superior effect on inhibiting ice recrystallization compared with WBAP with low A/X. WBAP could significantly improve the quality of frozen dough steamed bread, and the effect coincided with antifreeze activity. WBAP effectively inhibited the ice recrystallization and altered the shape of ice crystal from cubic to lamellate in frozen dough, consequently better preserving the yeast viability and dough structure. WBAP alleviated the depolymerization of glutenin macropolymers, and was conductive in developing more homogeneous and less distorted gluten network structures in frozen dough. The current study can provide a theoretical basis for the exploitation of WBAP as a technofunctional improver for frozen dough.

Interactomes in the era of deep learning.

Deep learning provides an atomic snapshot of the yeast protein interactome.

Forced association of SARS-CoV-2 proteins with the yeast proteome perturb vesicle trafficking.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the highly infectious coronavirus disease COVID-19. Extensive research has been performed in recent months to better understand how SARS-CoV-2 infects and manipulates its host to identify potential drug targets and support patient recovery from COVID-19. However, the function of many SARS-CoV-2 proteins remains uncharacterised. Here we used the Synthetic Physical Interactions (SPI) method to recruit SARS-CoV-2 proteins to most of the budding yeast proteome to identify conserved pathways which are affected by SARS-CoV-2 proteins. The set of yeast proteins that result in growth defects when associated with the viral proteins have homologous functions that overlap those identified in studies performed in mammalian cells. Specifically, we were able to show that recruiting the SARS-CoV-2 NSP1 protein to HOPS, a vesicle-docking complex, is sufficient to perturb membrane trafficking in yeast consistent with the hijacking of the endoplasmic-reticulum–Golgi intermediate compartment trafficking pathway during viral infection of mammalian cells. These data demonstrate that the yeast SPI method is a rapid way to identify potential functions of ectopic viral proteins.

Ubiquitin Ligase Redundancy and Nuclear-Cytoplasmic Localization in Yeast Protein Quality Control.

The diverse functions of proteins depend on their proper three-dimensional folding and assembly. Misfolded cellular proteins can potentially harm cells by forming aggregates in their resident compartments that can interfere with vital cellular processes or sequester important factors. Protein quality control (PQC) pathways are responsible for the repair or destruction of these abnormal proteins. Most commonly, the ubiquitin-proteasome system (UPS) is employed to recognize and degrade those proteins that cannot be refolded by molecular chaperones. Misfolded substrates are ubiquitylated by a subset of ubiquitin ligases (also called E3s) that operate in different cellular compartments. Recent research in Saccharomyces cerevisiae has shown that the most prominent ligases mediating cytoplasmic and nuclear PQC have overlapping yet distinct substrate specificities. Many substrates have been characterized that can be targeted by more than one ubiquitin ligase depending on their localization, and cytoplasmic PQC substrates can be directed to the nucleus for ubiquitylation and degradation. Here, we review some of the major yeast PQC ubiquitin ligases operating in the nucleus and cytoplasm, as well as current evidence indicating how these ligases can often function redundantly toward substrates in these compartments.