Higher Protein Yield Research Articles

Identification of potential candidate genes associated with milk protein differences in Holstein cows: a meta-analysis integrating GWAS and RNA-Seq transcriptome

Despite the identification of candidate genes influencing milk protein, the connections between genes and regulatory pathways remains elusive. This study aimed integrate findings from genome-wide association studies (GWAS) and RNA sequencing (RNA-Seq) through meta-analysis to pinpoint single nucleotide polymorphisms (SNPs) and genes responsible for high and low protein yield in cows. Previous GWAS and RNA-Seq analyses had identified 663 SNPs and 1106 genes ( P < 0.05). Twenty SNPs from GWAS, 10 genes from RNA-Seq, and 49 SNP/gene associations from both datasets, were identified using meta-analysis. Meta-analysis validated several SNPs previously identified through GWAS, such as rs135549651 ( P = 2.6 × 10−256), rs109146371 ( P = 3.1 × 10−208), rs109350371 ( P = 4.0 × 10−207), and rs109774038 ( P = 8.6 × 10−587). Genes identified in RNA-Seq experiments, including NR4A1 ( P = 3.2 × 10−7), ATF3 ( P = 9.6 × 10−7), CDH16 ( P = 9.9 × 10−7), VEGFA ( P = 1.0 × 10−6), and SAA3 ( P = 7.3 × 10−11), were confirmed. The combined GWAS and RNA-Seq datasets highlighted CCND2 ( P = 8.9 × 10−111), MAPK15 ( P = 1.3 × 10−151), and CPSF1 ( P = 1.2 × 10−306) as the most significant genes. Additionally, significant gene ontology (GO) terms, including ionizing radiation ( P = 1.5 × 10−4), nuclear pore cytoplasmic filaments ( P = 9.4 × 10−5), and phenylalanine 4-monooxygenase activity ( P = 1.4 × 10−5), were identified. In conclusion, the integration of GWAS and RNA-Seq, coupled with GO enrichment, allowed identification of candidate SNPs and genes with higher accuracy. These findings improve our knowledge about genomic architecture of milk protein and enhance evaluation of Holstein cows.

Bacillus cereus-mediated biofermentation of Sardine offal waste: A novel approach to enhance nutritional value by Response Surface Methodology optimization

The rising protein demand in the aquaculture sector has significantly impacted fishmeal supply and pricing. Excessive use of fishmeal can lead to environmental issues and negatively impact marine biodiversity and human food security. Consequently, finding alternative fishmeal in aquaculture is crucial for economic and environmental sustainability. The present study aimed to determine how Bacillus cereus (MT355408) could enhance nutritional value of Sardine fish waste, which could replace fish meal in the market. Solid-state fermentation (SSF) represents a biotechnological method that utilizes microbes to convert discarded fish byproducts into valuable products. The bacterial ability to produce enzymes was studied and optimised for its maximum production to be used as an inoculum for the SSF technique. Different prebiotic sources were also studied for better upliftment of bacteria in the solid-state surface. A single-factor analysis was conducted to investigate the influence of varying prebiotic concentrations, inoculum quantity, and fermentation duration on protein breakdown. After studying the single-factor tests, a further response surface model was employed for better yield. The results indicated that the highest protein yield could be achieved with a fermentation time of 132.893 hours, a prebiotic quantity of 25%, and an inoculum quantity of 5.3%. The study's findings also affirmed that the model was vital in enhancing the crude protein content during fermentation. In conclusion, the model's results contribute valuable insights into fermentation processes, offering practical implications for enhancing protein content and digestibility in similar contexts.

Transport-controlled growth decoupling for self-induced protein expression with a glycerol-repressible genetic circuit.

Decoupling cell formation from recombinant protein synthesis is a potent strategy to intensify bioprocesses. Escherichia coli strains with mutations in the glucose uptake components lack catabolite repression, display low growth rate, no overflow metabolism, and high recombinant protein yields. Fast growth rates were promoted by the simultaneous consumption of glucose and glycerol, and this was followed by a phase of slow growth, when only glucose remained in the medium. A glycerol-repressible genetic circuit was designed to autonomously induce recombinant protein expression. The engineered strain bearing the genetic circuit was cultured in 3.9 g L-1 glycerol + 18 g L-1 glucose in microbioreactors with online oxygen transfer rate monitoring. The growth was fast during the simultaneous consumption of both carbon sources (C-sources), while expression of the recombinant protein was low. When glycerol was depleted, the growth rate decreased, and the specific fluorescence reached values 17% higher than those obtained with a strong constitutive promoter. Despite the relatively high amount of C-sourceused, no oxygen limitation was observed. The proposed approach eliminates the need for the substrate feeding or inducers addition and is set as a simple batch culture while mimicking fed-batch performance.

Synergistic promotion of transient transgene expression in CHO cells by PDI/XBP-1s co-transfection and mild hypothermia.

Transient gene expression system is an important tool for rapid production of recombinant proteins in Chinese hamster ovary (CHO) cells. However, their low productivity is the main hurdle to overcome. An effective approach through which to obtain high protein yield involves targeting transcriptional, post-transcriptional events (PTEs), and culture conditions. Here, we investigated the effects of protein disulfide isomerase (PDI) and spliced X-box binding protein 1 (XBP-1s) co-overexpression combined with mild hypothermia on the transient yields of recombinant proteins in CHO cells. The results showed that the gene of interest (GOI) and the PDI/XBP-1s helper vector at a co-transfection ratio of 10:1 could obviously increase transient expression level of recombinant protein in CHO cells. However, PDI/XBP-1s overexpression had no significance effect on the mRNA levels of the recombinant protein, suggesting that it targeted PTEs. Moreover, the increased production was due to the enhancing of cell specific productivity, not related to cell growth, viability, and cell cycle. In addition, combined PDI/XBP-1s co-overexpression and mild hypothermia could further improve Adalimumab expression, compared to the control/37°C and PDI/XBP-1s/37°C, the Adalimumab volume yield of PDI/XBP-1s/33°C increased by 203% and 142%, respectively. Mild hypothermia resulted in 3.52- and 2.33-fold increase in the relative mRNA levels of PDI and XBP-1s, respectively. In conclusion, the combination of PDI/XBP-1s overexpression and culture temperature optimization can achieve higher transient expression of recombinant protein, which provides a synergetic strategy to improve transient production of recombinant protein in CHO cells.

Using Vibrio natriegens for High-Yield Production of Challenging Expression Targets and for Protein Perdeuteration.

Production of soluble proteins is essential for structure/function studies; however, this usually requires milligram amounts of protein, which can be difficult to obtain with traditional expression systems. Recently, the Gram-negative bacterium Vibrio natriegens emerged as a novel and alternative host platform for production of proteins in high yields. Here, we used a commercial strain derived from V. natriegens (Vmax X2) to produce soluble bacterial and fungal proteins in milligram scale, which we struggled to achieve in Escherichia coli. These proteins include the cholera toxin (CT) and N-acetyl glucosamine-binding protein A (GbpA) from Vibrio cholerae, the heat-labile enterotoxin (LT) from E. coli and the fungal nematotoxin CCTX2 from Coprinopsis cinerea. CT, GbpA, and LT are secreted by the Type II secretion system in their natural hosts. When these three proteins were produced in Vmax, they were also secreted and could be recovered from the growth media. This simplified the downstream purification procedure and resulted in considerably higher protein yields compared to production in E. coli (6- to 26-fold increase). We also tested Vmax for protein perdeuteration using deuterated minimal media with deuterium oxide as solvent and achieved a 3-fold increase in yield compared to the equivalent protocol in E. coli. This is good news, since isotopic labeling is expensive and often ineffective but represents a necessary prerequisite for some structural biology techniques. Thus, Vmax represents a promising host for production of challenging expression targets and for protein perdeuteration in amounts suitable for structural biology studies.

Komagataella phaffii as a Platform for Heterologous Expression of Enzymes Used for Industry.

In the 1980s, Escherichia coli was the preferred host for heterologous protein expression owing to its capacity for rapid growth in complex media; well-studied genetics; rapid and direct transformation with foreign DNA; and easily scalable fermentation. Despite the relative ease of use of E. coli for achieving the high expression of many recombinant proteins, for some proteins, e.g., membrane proteins or proteins of eukaryotic origin, this approach can be rather ineffective. Another microorganism long-used and popular as an expression system is baker's yeast, Saccharomyces cerevisiae. In spite of a number of obvious advantages of these yeasts as host cells, there are some limitations on their use as expression systems, for example, inefficient secretion, misfolding, hyperglycosylation, and aberrant proteolytic processing of proteins. Over the past decade, nontraditional yeast species have been adapted to the role of alternative hosts for the production of recombinant proteins, e.g., Komagataella phaffii, Yarrowia lipolytica, and Schizosaccharomyces pombe. These yeast species' several physiological characteristics (that are different from those of S. cerevisiae), such as faster growth on cheap carbon sources and higher secretion capacity, make them practical alternative hosts for biotechnological purposes. Currently, the K. phaffii-based expression system is one of the most popular for the production of heterologous proteins. Along with the low secretion of endogenous proteins, K. phaffii efficiently produces and secretes heterologous proteins in high yields, thereby reducing the cost of purifying the latter. This review will discuss practical approaches and technological solutions for the efficient expression of recombinant proteins in K. phaffii, mainly based on the example of enzymes used for the feed industry.

Impacts of Harvest Date and Concurrent Alkali Pretreatment and Ensiling on Anaerobic Digestion of Pennycress Biomass

Pennycress (Thlaspi arvense L.) is an annual cover crop known for its exceptional cold tolerance and high oil and protein yields. Pennycress can be integrated into a corn–soybean rotation in the U.S. However, the utilization of pennycress biomass remains largely unexplored, including assessing compositional changes through its growth and organic matter digestibility. This study harvested pennycress at three growth stages, characterized the biomass for anaerobic digestion (AD), and tested the effects of concurrent alkali pretreatment and ensiling on the biomass methane yield. Results showed that the biomass harvested when the plants were undergoing senescence (“third-harvest”) had higher contents of acid detergent fiber, neutral detergent fiber, and lignin, while the biomass harvested when 80–90% of the pods were fully-sized (“second-harvest”) had the highest protein content. The AD experiments showed that the first-harvest biomass (90% of flowers opened) failed to produce biogas due to a drop in the pH and alkalinity, the second-harvest biomass was inhibited for methane production (45.74 ± 0.20 L/kg-VS), and the third-harvest biomass had a methane yield of 171.80 ± 4.82 L/kg-VS. After the alkali pretreatment and ensiling, a methane yield of 270.4 ± 3.10 L/kg-VS was obtained from the second-harvest biomass, representing a significant 4.5-fold increase (adjusted for the organic matter loss) relative to the untreated second-harvest biomass.

Продуктивность коров при разной комбинации линий

The purpose of the study was to study the milk productivity of cows with different combinations of lines. The object of the study were first-calf cows of the black-and-white breed of the line Vis Back Aidiala 101341, Reflection Soveringa 198998, Montvik Chiefteina 95679. Cows obtained by intraline selection of the Aidial line have a high level of milk productivity, with milk yield of 5571 kg of milk, milk fat yield of 226 kg and protein yield of 171 kg, while significantly (P<0.05-0.01) superior to first-calf heifers, belonging to the Sovering line for 102 kg, 4 kg of fat and 4 kg of protein, and to the Chieftain line - for 259 kg of milk, 16 kg of fat and 11 kg of protein, respectively. When crossing the Ideal-Sovering lines, the highest milk yield is observed (5667 kg), with good milk fat content (4.01 %), high yield of milk fat (227 kg) and milk protein (172 kg). With the reverse cross Sovering-Idial, slightly lower milk productivity rates were obtained. When using bulls of the Aidial line on cows of the Governer line, high milk fat content (4.07 %) and milk fat yield (228 kg) were obtained, with average values of milk yield (5596 kg), protein mass fraction (3.01 %) and its yield (168 kg). Unsuccessful combinations of lines in both forward and reverse directions are crosses of the Idial and Chieftain lines. The descendants of such combinations have the lowest milk productivity. Thus, the use of line manufacturers R. Sovering and V.B. Idiala, both with intraline selection and interline combination, gives the best milk productivity indicators. Consequently, it is necessary to evaluate bulls according to the compatibility of lines; this has a great influence on the productivity of the offspring, since sires of the same line, when inseminating cows of the same lines, produce offspring of different productivity.

Strategies to Improve Quality of Fibrous Feeds: Evaluating the potential of Cow Pea (Vigna Unguiculata) accessions grown under sub-humid climatic conditions of Western Ethiopia

Five selected cowpea accessions (ILRI 9325, ILRI 11976, ILRI 6782, ILRI 6783 and WWT) were evaluated for herbage dry matter (DMY) and crude protein (CPY) yields, and nutritive value. Among the tested accessions, ILRI 9325, ILRI 6782, ILRI 6783 and WWT gave higher and comparable yields (P<0.01 for DMY; P=0.05 for CPY), with ILRI 9325 giving superior yields. The overall CP content was 17% with a range of 16% for WWT to 18% for ILRI 9325. The overall mean for neutral detergent fiber (NDF), acid detergent fiber (ADF) and lignin were 43%, 34% and 6%, respectively. The in vitro organic digestibility (IVOMD) ranged from 64% (ILRI 6783) to 69% (WWT). The relative feed value (RFV) index averaged 135 (range = 128 – 142). The mean metabolizable energy (ME) content was 10 MJ/kg DM with a range of 9.5 to 10.4. Three accessions that combined high biomass DM and crude protein yields, and herbage quality were: ILRI 9325, ILRI 11976 and WWT. The two former accessions can thus be promoted to the next variety verification stage alongside with WWT as a check to identify one or two alternative cowpea varieties for official release.

High throughput parameter estimation and uncertainty analysis applied to the production of mycoprotein from synthetic lignocellulosic hydrolysates

The current global food system produces substantial waste and carbon emissions while exacerbating the effects of global hunger and protein deficiency. This study aims to address these challenges by exploring the use of lignocellulosic agricultural residues as feedstocks for microbial protein fermentation, focusing on Fusarium venenatum A3/5, a mycelial strain known for its high protein yield and nutritional quality. We propose a high throughput microlitre batch fermentation system paired with analytical chemistry to generate time series data of microbial growth and substrate utilisation. An unstructured biokinetic model was developed using a bootstrap sampling approach to quantify uncertainty in the parameter estimates. The model was validated against an independent data set of a different glucose-xylose composition to assess the predictive performance. Our results indicate a robust model fit with high coefficients of determination and low root mean squared errors for biomass, glucose, and xylose concentrations. Estimated parameter values provided insights into the resource utilisation strategies of Fusarium venenatum A3/5 in mixed substrate cultures, aligning well with previous research findings. Significant correlations between estimated parameters were observed, highlighting challenges in parameter identifiability. The high throughput workflow presents a novel, rapid methodology for biokinetic model development, enabling efficient exploration of microbial growth dynamics and substrate utilisation. This innovative method directly supports the development of a foundational model for optimising microbial protein production from lignocellulosic hydrolysates, contributing to a more sustainable global food system.

БЕЛКОВАЯ ПРОДУКТИВНОСТь РАННЕСПЕЛЫХ СОРТОВ СОИ В УСЛОВИЯХ КАЛУЖСКОЙ ОБЛАСТИ

The article presents data on the duration of the growing season, biomass accumulation, seed productivity, protein content in seeds and protein yield with seed yield of four earlymaturing soybean varieties – Magheva, Kasatka, Svetlaya, and Georgiya of the selection of the Institute of Seed Production and Agrotechnology – a branch of the Federal Scientific Agroengineering Center VIM. The study of early-maturing soybean varieties capable of sustainable maturing in the Nonchernozem zone is aimed at solving the problem of protein deficiency in grain crops produced in agricultural enterprises, which is important for the devel- opment of animal husbandry. The research was carried out in 2018–2019 on sod-podzolic, sandy loam soils. The Svetlaya and Kasatka soybean varieties had the shortest growing season – 94 days, the Magheva variety had a 9-day longer growing season and the Georgiya variety had a 14-day longer growing season. The earliest maturing soybean varieties are Svet- laya and Kasatka. The sum of temperatures at the level of 1623°C is sufficient for the formation of seed yield. The soybean varieties Magheva and Georgiya require a sum of temperatures 150–200°C higher. The varieties with a long growing season – Georgiya and Magheva – are characterized by a high level of biomass accumulation and higher seed yield, on average over two years their seed yields were 1.64 and 1.48 t/ha respectively, while those of the varieties Svetlaya and Kasatka were 1.30 and 1.34 t/ha respectively. The soybean variet- ies Magheva (40.7%) and Kasatka (39.8%), while Georgiya (36.1%) had the lowest. Protein yield with the seed yield was more dependent on the seed yield. The correlation coefficient between protein yield and seed yield was 0.91, and between protein yield and seed protein content was 0.19. The varieties with the highest protein yields were Magheva and Georgiya with 602 and 599 kg/ha respectively. The varieties with the lowest protein yield were Kasatka (529 kg/ha) and Svetlaya (500 kg/ha).

Assessing the Impact of N Fertilizer on Soybean Yield and Quality Using a Modeling Approach

HighlightsSimulating the effect of high N fertilizer rates with the CSM-CROPGRO-Soybean model for Brazil.Simulated and observed plant tissue N accumulation increased with an increase in N fertilizer rates.N use efficiency decreased with N fertilization on soybeans under long-term scenarios.Both model simulations and observations indicate only a small response in crop yield to high N fertilization.Abstract. Soybean [Glycine max (L.)] is one of the main sources of food protein in the world. To achieve a high yield and grain protein concentration, soybean requires a large amount of nitrogen (N). Many studies have evaluated N fertilization in soybean, with the hypothesis that symbiotic N fixation may not be sufficient to supply N for high-yielding soybean. This study aimed to evaluate the performance of the CSM-CROPGRO-Soybean model with field data to determine the impact of N fertilizer on soybean and explore long-term scenarios. The experiments consisted of a combination of available water management and N fertilizer levels over eight crop seasons. After calibration and evaluation, the CSM-CROPGRO-Soybean model was used to explore the soil and plant N balance. We also applied the model to explore different N fertilization rates in long-term scenarios. The results showed that the model was able to simulate soybean growth with a very good agreement for all sites (D-statistic ranging from 0.63 to 0.99). The observed and simulated crop yield gains from N fertilization were small and not proportional to the amount of N fertilizer applied. The model underestimated the effect of N fertilizer on grain protein concentration increment, but there was a consistent trend towards an increase in protein concentration observed in measured data. Simulations of soil inorganic N balance suggested very high losses of N by volatilization for urea-N fertilization treatments. The long-term scenarios showed that using N rates above 600 kg ha-1 is potentially inappropriate, and a higher crop N use efficiency was obtained with no N fertilizer. Keywords: Ammonia volatilization, Crop yield, Crop N use efficiency, CROPGRO-Soybean model, Grain protein concentration, Nitrogen balance.

Removal of organic matter during adaptation of Nannochloropsis oculata in livestock waste

Microalgae, like plants, contribute significantly to the development of the oxygen biogeochemical cycle due to their high photosynthetic efficiency. In addition, they provide high yields of polyunsaturated fatty acids, sterols, proteins, terpenoids, and pigments, among others. Therefore, different species of microalgae have been studied and used on a laboratory scale to carry out processes such as wastewater treatment or aerobic bioconversion, which are presented as sustainable and viable alternatives for the treatment and recovery of organic waste (OW), usually rich in carbohydrates, lipids and proteins. In the present investigation, the removal of organic matter was evaluated during the adaptation of Nannochloropsis oculata in residues of poultry wastewater and swine origin, obtained from technical and semi-technical plants, respectively, located in the high mountain zone of the state of Veracruz, Mexico. The experiment was carried out in 250 mL discontinuous photobioreactors with a working volume of 200 mL, where 3 inoculum-substrate ratios were studied for each organic residue: 10, 15 and 20% inoculum in poultry wastewater (PWW) and 30, 50 and 70% inoculum in pig manure (PM). In addition, the conditions of temperature (20 ± 2 °C), illumination (2000 lx), photoperiod of 12/12 (light/dark) and continuous aeration were controlled. It was shown that N. oculata can tolerate alkaline conditions of pH ≥ 10 and contributes to the reduction of soluble organic matter in OW. PWW and PM were found to be viable media for the survival of N. oculata. Finally, regarding the inoculum concentrations studied, the most appropriate were 10% for PWW and 70% for PM.

Determination of process parameters and precipitation methods for potential large-scale production of sugar beet leaf protein concentrate.

Sugar beet is one of the most produced industrial plants in the world, and during manufacturing it produces a large quantity of leaf waste. Because this waste is rich in protein, this study aimed to identify an efficient method for producing large-scale protein concentrate from sugar beet leaves. Results showed that protein extraction from fresh leaves was more effective than from dried leaves. Maximum protein extraction was achieved at pH 9, compared with pH 7 or 8. Blanching as a pretreatment reduced protein yield during isoelectric precipitation, with a yield of 2.31% compared to 20.20% without blanching. Consequently, blanching was excluded from the extraction process. After extraction, isoelectric precipitation, heat coagulation, and isoelectric-ammonium sulfate precipitation were compared. Although the latter resulted in the highest protein yield, Fourier transform infrared analysis revealed that excessive salt was not removed during dialysis, making it unsuitable for scale-up due to its additional cost and complexity. Therefore, isoelectric precipitation was selected as the appropriate method for protein precipitation from sugar beet leaves. To increase yield, extractions were assisted by ultrasound or enzyme addition. Ultrasound-assisted extraction resulted in an increased protein yield from 20.20% to 28.60%, while Pectinex Ultra SP-L-assisted extraction was the most effective, increasing protein yield from 20.20% to 38.09%. Proteins were extracted from fresh sugar beet leaves using optimum conditions (50 °C, 30 min, pH 9) and precipitated at isoelectric point, with enzymatic-assisted extraction yielding the maximum protein recovery. © 2023 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Introgression of a Danbaekkong high-protein allele across different genetic backgrounds in soybean.

Soybean meal is a major component of livestock feed due to its high content and quality of protein. Understanding the genetic control of protein is essential to develop new cultivars with improved meal protein. Previously, a genomic region on chromosome 20 significantly associated with elevated protein content was identified in the cultivar Danbaekkong. The present research aimed to introgress the Danbaekkong high-protein allele into elite lines with different genetic backgrounds by developing and deploying robust DNA markers. A multiparent population consisting of 10 F5-derived populations with a total of 1,115 recombinant inbred lines (RILs) was developed using "Benning HP" as the donor parent of the Danbaekkong high-protein allele. A new functional marker targeting the 321-bp insertion in the gene Glyma.20g085100 was developed and used to track the Danbaekkong high-protein allele across the different populations and enable assessment of its effect and stability. Across all populations, the high-protein allele consistently increased the content, with an increase of 3.3% in seed protein. A total of 103 RILs were selected from the multiparent population for yield testing in five environments to assess the impact of the high-protein allele on yield and to enable the selection of new breeding lines with high protein and high yield. The results indicated that the high-protein allele impacts yield negatively in general; however, it is possible to select high-yielding lines with high protein content. An analysis of inheritance of the Chr 20 high-protein allele in Danbaekkong indicated that it originated from a Glycine soja line (PI 163453) and is the same as other G. soja lines studied. A survey of the distribution of the allele across 79 G. soja accessions and 35 Glycine max ancestors of North American soybean cultivars showed that the high-protein allele is present in all G. soja lines evaluated but not in any of the 35 North American soybean ancestors. These results demonstrate that G. soja accessions are a valuable source of favorable alleles for improvement of protein composition.

A nanobody-based strategy for rapid and scalable purification of human protein complexes.

The isolation of proteins in high yield and purity is a major bottleneck for the analysis of their three-dimensional structure, function and interactome. Here, we present a streamlined workflow for the rapid production of proteins or protein complexes using lentiviral transduction of human suspension cells, combined with highly specific nanobody-mediated purification and proteolytic elution. Application of the method requires prior generation of a plasmid coding for a protein of interest (POI) fused to an N- or C-terminal GFP or ALFA peptide tag using a lentiviral plasmid toolkit we have designed. The plasmid is then used to generate human suspension cell lines stably expressing the tagged fusion protein by lentiviral transduction. By leveraging the picomolar affinity of the GFP and ALFA nanobodies for their respective tags, the POI can be specifically captured from the resulting cell lysate even when expressed at low levels and under a variety of conditions, including detergents and mild denaturants. Finally, rapid and specific elution of the POI (in its tagged or untagged form) under native conditions is achieved within minutes at 4 °C, using the engineered SUMO protease SENPEuB. We demonstrate the wide applicability of the method by purifying multiple challenging soluble and membrane protein complexes to high purity from human cells. Our strategy is also directly compatible with many widely used GFP-expression plasmids, cell lines and transgenic model organisms. Finally, our method is faster than alternative approaches, requiring only 8 d from plasmid to purified protein, and results in substantially improved yields and purity.

High‐pressure‐assisted protein isolation from pumpkin seed press cake

SummaryTraditional solvent‐based approaches for the separation of protein from plant sources are widely used, although they can lead to low protein yield and protein degradation. In this study, protein isolation was conducted by alkaline extraction (AE) and isoelectric precipitation, and the effect of high‐pressure homogenisation (HPH) (0 MPa [control], 50, 100, and 150 MPa) at two different stages during extraction was assessed, namely high pressure homogenisation followed by alkaline extraction (HPH + AE) or alkaline extraction followed by high pressure homogenisation (AE + HPH). Although the control procedure resulted in a protein yield of 45.99 ± 0.72%, the highest protein yield was determined by AE + 100 MPa HPH (61.92 ± 1.06%). Solubility, foaming capacity, emulsification index, hue angle, and denaturation enthalpy of the isolated protein increased after HPH treatments compared to control. Generally, the high‐pressure treatment during protein isolation increased pumpkin seed press cake's protein yield and enhanced the isolates' techno‐functional properties.

Potential of enriched phototrophic purple bacteria for H2 bioconversion into single cell protein

Single cell protein (SCP) has emerged as an alternative protein source, potentially based on the recovery of carbon and nutrients from waste-derived resources as part of the circular economy. From those resources, gaseous substrates have the advantage of an easy sterilization, allowing the production of pathogen-free SCP. Sterile gaseous substrates allow producing pathogen-free SCP. This study evaluated the use of an enriched phototrophic purple bacteria (PPB) consortium for SCP production using H2 and CO2 as electron and C sources. The influence of pH (6.0–8.5), temperature (15–50 °C) and light intensity (0–50 W·m−2) on the growth kinetics and biomass yields was investigated using batch tests. Optimal conditions were found at pH 7, 25 °C and light intensities over 30 W·m−2. High biomass and protein yields were achieved (~ 1 g CODbiomass·g CODH2consumed−1 and 3.9–4.4 g protein·g H2−1) regardless of the environmental conditions, being amongst the highest values reported from gaseous streams. These high yields were obtained thanks to the use of light as a sole energy source by the PPB consortium, allowing a total utilization of H2 for growth. Hydrogen uptake rates varied considerably, with values up to 61 ± 5 mg COD·d−1 for the overall H2 consumption rates and 2.00 ± 0.14 g COD·g COD−1·d−1 for the maximum specific uptake rates under optimal growth conditions. The latter value was estimated using a mechanistic model able to represent PPB growth on H2. The biomass exhibited high protein contents (>50 % w/w) and adequate amino acid profiles, showing its suitability as SCP for feed. PPB were the dominant bacteria during the experiments (relative abundance over 80 % in most tests), with a stable population dominated by Rhodobacter sp. and Rhodopseudomonas sp. This study demonstrates the potential of enriched PPB cultures for H2 bioconversion into SCP.

Influence of weather conditions on yield and protein content in winter wheat grain

The results of three-year studies (2020–2022) conducted in the forest-steppe zone of the Samara region in the agrocenosis of winter wheat are presented. The purpose of the work was to study the influence of natural and climatic conditions of the research area on the formation of yield indicators, protein content in winter wheat grain. The material was 10 varieties from the Vavilov All-Russian Institute of Plant Genetic Resources (VIR) collection and 5 varieties of selection of the Povolzhsky Scientific Research Institute of Selection and Seed Growing named after P.N. Konstantinov. Yield, grain protein content and protein yield per hectare were determined. Varieties characterized by high yield and high amount of protein in grain, showing maximum protein yield in different weather conditions of the years were identified. Over the years of research, the year 2022 was the most favorable for the formation of high yields (between 6 and 8.5 t/ha were harvested in the indicated year), where a large amount of precipitation was observed in May – the first ten-day period of June. The protein content in grain was the highest in 2021 (14–19%), when high temperatures were observed during the grain filling period in the absence of precipitation. For each variety, the correlation dependence of the sum of active temperatures and the amount of precipitation during the spring-summer vegetation period with the studied indicators was established. There was a negative correlation between the sum of active temperatures and yield (from –0.553 to –0.981) and a positive correlation with the protein content indicator (from 0.605 to 0.984) depending on the variety. The amount of precipitation was in a positive correlation with the yield index, the dependence on the protein content in the grain was ambiguous and depended on the variety. The studied indicators were influenced not only by natural and climatic conditions, but also by the genotypic features of the studied varieties, which explains the diverse correlation dependence.

Fractionation of proteins from alfalfa juice at demonstration scale: Effects of temperature and centrifugation strategies on removal of green protein and recovery of the soluble white protein fraction

Given the growing global demand for protein, alfalfa (Medicago sativa) has emerged as a promising alternative protein source for both animal feed and human nutrition due to its high protein content and nutritional value. This study investigated the fractionation of proteins in freshly pressed alfalfa juice into green and white fractions, with focus on optimizing the recovery of the white protein fraction, which holds significant potential for various food applications. Two sets of demonstration-scale experiments were conducted to evaluate the impact of temperature and centrifugation techniques on the fractionation process. The goal was to achieve the optimal coagulation and removal of the green fraction while concurrently enhancing the yield of the white protein fraction. The first set of experiments explored heat treatment at four different temperatures (50 °C, 55 °C, 60 °C, and 65 °C), followed by decanter centrifugation. Optimal coagulation of the green fraction with higher white protein fraction yield was observed at 55 °C. In the second set of experiments, a constant temperature was maintained (55 °C) while testing three different separation scenarios. A two-stage centrifugation approach employing decanter and disc-stack high-speed centrifuges proved most effective, removing most of the coagulated green fraction and preserving over 80% of the white fraction in the residual juice after heat treatment. Complete removal of the green fraction was not achieved, and an additional microfiltration step was introduced to clarify the residual juice. Microfiltration retained up to 50 % of the white protein, resulting in a significant reduction in the overall recovery yield. The results indicate that achieving high recovery yields of soluble white protein remains a challenge and requires further optimization of the fractionation process.