Biology subjects

Gregersen Echers, S.

Publications and source records attributed to Gregersen Echers, S..

9 recordsLinked to original sources

bioRxiv2026

Protein concentrates from pilot-scale biorefining of clover grass: Interfacial properties and their effects on the physical and oxidative stability of fish oil-in-water emulsion

Clover grass blends are promising sources of nutritional and techno-functional proteins, but currently mainly utilized for animal feeding. The application as a physical and oxidative stabilizer in food emulsions remains underexplored. In this study, the stabilizing effects of clover grass proteins (CGPs), produced through a pilot-scale, two-stage membrane filtration process yielding a native GPC concentrate (DC), as well as enzymatic hydrolysate hereof (DCH), were compared with commercial plant proteins (soy and pea) and animal sources (sodium caseinate). Both DC and DCH produced emulsions (0.4% (w/w) protein and 5% fish oil) with smaller size droplets and larger electrostatic repulsion between droplets compared to the other proteins tested. Moreover, DC and DCH exhibited higher protection against the generation of both primary and secondary oxidation products. Furthermore, emulsions stabilized with CGPs were well-protected from off-flavor compounds. Mass spectrometry-based proteomics analysis revealed that DC included a high RuBisCO content (38%) and the membrane process successfully depleted pigment-binding proteins affiliated with grassy color and sensory attributes. Moreover, DC was enriched (compared to the initial green juice) in known antioxidant proteins, constituting 10% of the total protein. In the hydrolysate (DCH), 30% of the total MS1 peptide signal originated from peptides predicted as probable free radical scavengers. These findings demonstrate that refined, native CGP, as well as its hydrolysate, improved both physical and oxidative stability of emulsions compared to plant and animal-based reference proteins due to a high endogenous antioxidant properties of the protein.

bioengineering↗

bioRxiv2026

Techno-functional properties of clover grass protein and the effect of different process technologies

This study investigated the effects of different downstream processes for protein isolation on the bulk properties and composition of clover grass protein prototypes (CGPs). A clarified clover grass juice, obtained using membrane filtration (MF), underwent precipitation by acid (AP), heat (HP), or acid+heat (AHP), or underwent ultra- and diafiltration to produce a concentrate (DC) as well as subsequent tryptic hydrolysis of DC (DCH). HP had the highest protein content (p<0.05) and was whiter than other CGPs, although it showed lower aqueous solubility. In contrast, DC showed excellent solubility across a broad pH range. CGPs efficiently decreased oil-water interfacial tension (16-13 mN/m) and displayed viscoelastic and solid-like interfacial behavior. CGPs-stabilized emulsions displayed low physical stability with larger droplets despite high absolute {zeta}-potentials. CGPs were rich in RuBisCO (37-47%) but had varying levels of other proteins. Despite significant protein-level differences, overall protein composition of CGPs was comparable, highlighting that protein state governs bulk functionality more than subtle compositional changes. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/701969v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@f37756org.highwire.dtl.DTLVardef@1fb5beorg.highwire.dtl.DTLVardef@1d4efe2org.highwire.dtl.DTLVardef@d11ef8_HPS_FORMAT_FIGEXP M_FIG C_FIG Created with BioRender.com HighlightsO_LIThe effect of different processes on functional properties of CGPs was explored. C_LIO_LIHeat treatment increased protein purity and whiteness at the expense of solubility. C_LIO_LICGPs efficiently reduced O/W interfacial tension but produced unstable emulsions. C_LIO_LICGPs were found rich in RuBisCO (34-47%) using quantitative proteomics. C_LIO_LIProtein state had larger influence on functionality than protein-level composition. C_LI

bioengineering↗

bioRxiv2025

Selective removal of green pigments and associated proteins from clover-grass protein concentrates: Molecular insight into a non-destructive, two-stage membrane-based biorefinery concept for high-quality food protein production

Green leaves are gaining traction as an emerging protein source and a sustainable alternative to animal-based protein as leaf proteins often possess good nutritional and functional properties. Current methods for producing protein isolates from leaves include partially or fully denaturing conditions, diminishing protein solubility and functionality. Here, we characterize the performance of a multi-stage, membrane-based green biorefinery concept capable of producing a native protein product from clover grasses devoid of attributes such as green color and grassy smell/taste. By sampling at each step along the process, we obtain insights on the fate of proteins and pigments over each unit operation. The process efficiently removes green pigments (>99.9%), when comparing the product stream with the initial feed stream, based on UV/Vis analysis. Using mass spectrometry-based proteomics, two complementary quantification strategies, and subsequent bioinformatic analysis, this can particularly be ascribed to a very selective retention (>99%) of unwanted membrane-associated and pigment-binding proteins in the first-stage filtration. In the second-stage filtration and subsequent diafiltration stage, residual unwanted proteins, fragments, and pigments are efficiently washed out while retaining the overall protein composition. The product maintains the high RuBisCO content of green juice and is furthermore enriched in proteins with known antioxidant properties while depleted in known food allergens. This work presents an in-depth understanding of protein-level selectivity in membrane-based green biorefinery and can help guide process optimization towards improved yields and quality. HighlightsO_LIStreams from a two-stage membrane-based green biorefinery were characterized C_LIO_LI>99.9% pigment-binding proteins were removed primarily in first stage filtration C_LIO_LIThe 2nd filtration and diafiltration removed residual pigment and retained protein C_LIO_LIThe full process efficiently removed >99.9% of chlorophylls and carotenoids C_LIO_LIThe soluble product is rich in RuBisCO and enriched in known antioxidant proteins C_LI

bioengineering↗

bioRxiv2025

Whats left from the brew? Investigating residual barley proteins in spent grains for downstream valorization opportunities

Brewers spent grain (BSG) is the major side-stream from beer production but remains highly underutilized. While the direct use of BSG as a food ingredient is limited due to subpar techno- functionality, the vast amounts and fairly high protein content of up to 30% makes it a high potential source for production of protein-based ingredient by valorization through e.g. enzymatic hydrolysis. However, little attention has been put towards the protein-level composition of BSG, which is essential for developing hydrolysis strategies for improving functionality in a targeted manner. Here, we present an in-depth characterization of the BSG proteome and investigate dynamic proteome changes from malting and mashing in the initial phases of beer production. We show dynamic and selective changes in the proteome across the different process steps, where 29% of reproducibly identified proteins display differential abundance. BSG represents a significantly higher proportion of intracellular protein compared to both barley and malt and has a nutritionally favorable amino acid composition. The major constituent of the BSG proteome is B3-Hordein, constituting more than 30% of the BSG protein. Moreover, we find that a large proportion (> 45%) of the BSG protein is associated with potential food safety concerns, being classified as potential allergens and antinutritional factors. Our analysis emphasizes the need for downstream processing of BSG to produce safe and functional food ingredients, while also providing protein-level insights for development of targeted hydrolysis strategies to achieve this. HighlightsO_LIAn optimized sample preparation for proteomics analysis has been developed C_LIO_LI29% of barley proteins are differentially abundant across malting and mashing C_LIO_LIB3-Hordein is the major protein in BSG with an abundance over 30% C_LIO_LIBSG contains a high content of potential allergenic and antinutritional proteins C_LIO_LIA protein-level basis for targeted downstream processing of BSG is presented C_LI

plant biology↗

bioRxiv2024

To fly, or not to fly, that is the question: A deep learning model for peptide detectability prediction in mass spectrometry

Identifying detectable peptides, known as flyers, is key in mass spectrometry-based proteomics. Peptide detectability is strongly related with the peptide sequence and its resulting physicochemical properties. Moreover, the high variability in MS data, particularly in peptide detectability and intensity across multiple analyses and samples, makes the development of a generic model for detectability prediction unfeasible. This underlines the need for tools that can be refined for specific experimental conditions. To address this need, we present Pfly, a deep learning model developed to predicts peptide detectability based solely on peptide sequence. Pfly distinguishes itself as a versatile and reliable state-of-the-art tool, offering high performance, accessibility, and easy customizability for end-users. This adaptability allows researchers to tailor the model to their specific experimental conditions, facilitating the creation of lab-specific models. This, in turn, can lead to more accurate results and expand the models applicability across various research fields. The models architecture is an encoder-decoder with an attention mechanism. This tool classifies peptides as either flyers or non-flyers, providing both binary probabilities and detailed categorical probabilities for four distinct classes defined in this study: non-flyer, weak flyer, intermediate flyer, and strong flyer. The model was initially trained on a synthetic peptide library and subsequently fine-tuned with a biological dataset to mitigate bias towards synthesizability, improving the predictive capacity and outperforming state-of-the-art predictors in a benchmark comparison. The study further investigates the influence of protein abundance and the search engine, illustrating the negative impact on peptide identification due to misclassification. Pfly has been integrated in the DLOmix framework and it is accessible on GitHub at https://github.com/wilhelm-lab/dlomix.

bioinformatics↗

bioRxiv2024

Decoding the Impact of Neighboring Amino Acid on MS Intensity Output through Deep Learning

Peptide-level quantification using mass spectrometry (MS) is no trivial task as the physicochemical properties affect both response and detectability. The specific amino acid (AA) sequence affects these properties, however the link from sequence to intensity output remains poorly understood. In this work, we explore combinations of amino acid pairs (i.e., dimer motifs) to determine a potential relationship between the local amino acid environment and MS1 intensity. For this purpose, a deep learning (DL) model, consisting of an encoder-decoder with an attention mechanism, was built. The attention mechanism allowed to identify the most relevant motifs. Specific patterns were consistently observed where a bulky/aromatic and hydrophobic AA followed by a cationic AA as well as consecutive bulky/aromatic and hydrophobic AAs were found important for the MS1 intensity. Correlating attention weights to mean MS1 intensities revealed that some important motifs, particularly containing Trp, His, and Cys, were linked with low responding peptides whereas motifs containing Lys and most bulky hydrophobic AAs were often associated with high responding peptides. Moreover, Asn-Gly was associated with low MS1 response. The model could predict MS1 response with a mean average percentage error of [~]11% and a Pearson correlation coefficient of [~]0.68.

bioinformatics↗

bioRxiv2023

Insight on physicochemical properties governing peptide MS1 response in HPLC-ESI-MS/MS proteomics: A deep learning approach

Accurate and absolute quantification of individual peptides in complex mixtures is a challenge not easily overcome. A potential solution is the use of quantitative mass spectrometry (MS) based methods, however, current state of the art requires foreground knowledge and isotopically labeled standards for each peptide to be accurately quantified. This increases analytical expenses, time consumption, and labor, limiting the number of peptides that can be quantified. A key step in developing less restrictive label-free quantitative peptidomics methods is understanding of the physicochemical properties of peptides that influence the MS response. In this work, a deep learning model was developed to identify the most relevant physicochemical properties based on repository MS data from equimolar peptide pools. Using an autoencoder with attention mechanism and correlating attention weights with corresponding physicochemical property indices from AAindex1, we were able to obtain insight on the properties governing the peptide-level MS1 response. These properties can be grouped in three main categories related to peptide hydrophobicity, charge, and structural propensities. Moreover, we present a model for predicting the MS1 intensity output based solely on peptide sequence input. Using a refined training dataset, the model predicted log-transformed peptide MS1 intensities with an average error of 11%.

bioinformatics↗

bioRxiv2022

Bioinformatically predicted emulsifying peptides and potato protein hydrolysate improves the oxidative stability of microencapsulated fish oil

The aim of this study was to investigate the potential of potato proteins and peptides as emulsifiers in the microencapsulation of fish oil by spray-drying. Microcapsules were produced using a potato protein extract, and fractions enriched in patatin and protease inhibitors. Furthermore, bioinformatically predicted emulsifier peptides from abundant potato proteins and a hydrolysate, obtained through targeted proteolysis of the extract, were investigated. During 28 days of storage at 25{degrees}C, peptides and hydrolysate exhibited better emulsifying properties and higher encapsulation efficiencies compared to native proteins and sodium caseinate. Significant differences (p < 0.05) were observed in the peroxide value (PV) and secondary volatile oxidation products between the microcapsules produced with peptides and native proteins. Microcapsules produced with peptides and hydrolysate showed the highest oxidative stability, not exceeding a PV of 10 meq/kg oil, and with concentrations of volatiles below the odor threshold in oil for five of the six studied compounds. These results show the emulsifying potential of potato peptides and hydrolysate for use in microencapsulation of hydrophobic bioactive ingredients such as fish oil.

bioengineering↗

bioRxiv2022

Targeted hydrolysis of native potato protein: A novel route for obtaining hydrolysates with improved interfacial properties

Peptides and protein hydrolysates are promising alternatives to substitute chemical additives as functional food ingredients. In this study, we present a novel approach for producing a potato protein hydrolysate with improved emulsifying and foaming properties by data-driven, targeted hydrolysis. Based on previous studies, we selected 15 emulsifier peptides derived from abundant potato proteins, which were clustered based on sequence identity. Through in silico analysis, we determined that from a range of industrial proteases (Neutrase (Neut), Alcalase (Alc), Flavorzyme (Flav) and Trypsin (Tryp)), Tryp was found more likely to release peptides resembling the target peptides. After applying all proteases individually, hydrolysates were assayed for in vitro emulsifying and foaming properties. No direct correlation between degree of hydrolysis and interfacial properties was found. Tryp produced a hydrolysate (DH=5.4%) with the highest (P<0.05) emulsifying and foaming abilities, good stabilities, and high aqueous solubility. Using LC-MS/MS, we identified >10,000 peptides in each hydrolysate. Through peptide mapping, we show that random overlapping with known peptide emulsifiers is not sufficient to quantitatively describe hydrolysate functionality. While Neut hydrolysates had the highest proportion of peptides with target overlap, they showed inferior interfacial activity. In contrast, Tryp was able to release specifically targeted peptides, explaining the high surface activity observed. While modest yields and residual unhydrolyzed protein indicate room for process improvement, this work shows that data-driven, targeted hydrolysis is a viable, interdisciplinary approach to facilitate hydrolysis design for production of functional hydrolysates from alternative protein sources.

biochemistry↗