Protein Secondary Structure Research Articles

T266M variants of ANGPTL4 improve lipid metabolism by modifying their binding affinity to acetyl-CoA carboxylase in obstructive sleep apnea

Background Angiopoietin-like protein 4 (ANGPTL4) is recognized as a crucial regulator in lipid metabolism. Acetyl-CoA carboxylases (ACACAs) play a role in the β-oxidation of fatty acids. Yet, the functions of ANGPTL4 and ACACA in dyslipidemia of obstructive sleep apnea (OSA) remain unclear. Methods This study included 125 male OSA subjects from the Shanghai Sleep Health Study (SSHS) who were matched for age, body mass index (BMI), and lipid profile. Serum ANGPTL4 levels were measured via ELISA. The ANGPTL4 T266M variants of 4455 subjects along with their anthropometric, fasting biochemical, and standard polysomnographic parameters were collected. Linear regression was used to analyze the associations between quantitative traits and ANGPTL4 T266M. Molecular docking and molecular dynamic simulation were employed to compare the effects of the wild-type ANGPTL4 and its T266M mutation on ACACA. Results Serum ANGPTL4 levels significantly decreased with increasing OSA severity (non-OSA: 59.6 ± 17.4 ng/mL, mild OSA: 50.0 ± 17.5 ng/mL, moderate OSA: 46.3 ± 15.5 ng/mL, severe OSA: 19.9 ± 14.3 ng/mL, respectively, p = 6.02 × 10−16). No associations were found between T266M and clinical characteristics. Molecular docking indicated that mutant ANGTPL4 T266M had stronger binding affinity for the ACACA protein, compared with wild-type ANGPTL4. In terms of protein secondary structure, mutant ANGTPL4 T266M demonstrated greater stability than wild-type ANGPTL4. Conclusions Serum ANGTPL4 levels were significantly decreased in OSA patients, particularly among individuals with severe OSA. Although functional ANGTPL4 T266M variants were not associated with lipid levels in OSA, ANGTPL4 T266M could enhance binding affinity for the ACACA protein, potentially regulating lipid metabolism.

Effect of hydrocavitation treatment on physicochemical and functional properties of soy protein isolate

A hydrocavitation reactor was employed to treat diluted samples of soy protein isolate (SPI) at room temperature for 0, 60, and 90-minute intervals, revealing significant improvements in vitro enzymatic digestion, trypsin inhibition, water holding, oil holding, emulsification, and solubility properties. The secondary protein structure underwent significant modifications, with an increase in surface area, etching phenomena, and structural aggregation. The extent of amino acid digestion for hydrocavitation-treated proteins exhibited an augmentation from 180.9 % ± 1.429 μg/mL to 189.5 % ± 0.556 μg/mL. Additionally, the water holding capacity of the treated proteins increased from 3.51 % ± 0.131 to 3.78 % ± 0.097, while there was a corresponding decrease in oil holding capacity. Solubility showed a noteworthy rise from 24.70 % ± 0.042 to 38.23 % ± 0.031 following the hydrocavitation treatment. These findings underscore the potential of hydrocavitation technology as an appealing avenue for non-thermal, chemical-free food processing, enhancing product quality and digestibility through minimal processing techniques.

Divalent metal ions under low concentration environment improved the thermal gel properties of egg yolk

To improve the thermal gel properties of egg yolk, the effect of several valence metal ions (K+, Ca2+, Mg2+ and Fe3+) with different concentrations (0-0.72%) on the rheological, gel, and structural properties of egg yolk were investigated. Results showed that monovalent and divalent ions were beneficial to the formation of uniform and dense gel network, especially with the addition of 0.72% magnesium ion, which further improved gel hardness, water holding capacity (WHC) and viscoelastic properties, the properties of egg yolk gel increased with the increase of the concentration of mono-bivalent metal ions. Adding ferric ion remarkably increased the average particle size (d4,3) and apparent viscosity of egg yolk, destroying the disulfide bonds and the hydrophobic interactions in gel. Fourier transform infrared spectroscopy (FT-IR) and fluorescence spectra analysis revealed that metal ions promoted the hydrophobic aggregation among egg yolk proteins and induced the transition of protein secondary structure from ordered to disordered. This work will provide a theoretical reference for the development of low salt and nutrient fortified egg yolk products.

Noninvasive evaluation of the skin barrier in reconstructed human epidermis using speckle analysis: Correlation with Raman microspectroscopy.

Reconstructed epidermis models, obtained from 3D keratinocytes culture, have gained significant prominence as prototypes for safety and efficacy testing in skin research. To effectively evaluate these models, it is essential to perform molecular and functional characterization. The skin's barrier function is one of the essential aspects of the epidermis that needs to be assessed. A noninvasive method is thus required for the evaluation of the skin barrier in these models. With this perspective, the aim of this feasibility study is to apply the speckle technique for the assessment of the skin barrier in the Reconstructed Human Epidermis (RHE). Speckle analysis as well as Raman microspectroscopy were performed on RHE samples at two maturation days, D17 and D20. Between D17 and D20, our study showed an increase in various Raman parameters, including stratum corneum percentage, lateral lipid packing, lipid-to-protein ratio, and protein secondary structure. Furthermore, the degree of light polarization and the speckle grain size also increased over this period. The speckle technique proved to be effective for evaluating the skin barrier in Reconstructed Human Epidermis (RHE) models. Comparison with Raman validates this approach and provides comprehensive molecular and functional characterization of reconstructive skin models.

Exploring the effect of protein secondary structure on the solid state and physical stability of protein-based amorphous solid dispersions

Amorphous solid dispersions (ASDs) using proteins as carriers have emerged as a promising strategy for stabilizing amorphous drug molecules. Proteins possess diverse three-dimensional structures that significantly influence their own properties and may also impact the properties of ASDs. We prepared β-lactoglobulin (BLG) with different contents of β-sheet and α-helical secondary structures by initially dissolving BLG in different mixed solvents, containing different ratios of water, methanol/ethanol, and acetic acid, followed by spray drying of the solutions. Our findings revealed that an increase in α-helical content resulted in a decrease in the glass transition temperature (Tg) of the protein. Subsequently, we utilized the corresponding mixed solvents to dissolve both BLG and the model drug celecoxib (CEL), allowing the preparation of ASDs containing either β-sheet-rich or α-helix/random coil-rich BLG. Using spray drying, we successfully developed BLG-based ASDs with drug loadings ranging from 10 wt% to 90 wt%. At drug loadings below 40 wt%, samples prepared using both methods exhibited single-phase ASDs. However, heterogeneous systems formed when the drug loading exceeded 40 wt%. At higher drug loadings, physical stability assessments demonstrated that the α-helix/random coil-rich BLG structure exerted a more pronounced stabilizing effect on the drug-rich phase compared to the β-sheet-rich BLG. Overall, our results highlight the importance of considering protein secondary structure in the design of ASDs.

An atomistic characterization of high-density lipoproteins and the conserved “LN” region of apoA-I

The physicochemical characteristics of the various subpopulations of high-density lipoproteins (HDLs) and, in particular, their surface properties determine their ability to scavenge lipids and interact with specific receptors and peptides. Five representative spheroidal HDL subpopulation models were mapped from a previously reported equilibrated coarse-grained (CG) description to an atomistic representation for subsequent molecular dynamics simulation. For each HDL model a range of finer-level analyses was undertaken, including the component-wise characterization of HDL surfaces, the average size and composition of hydrophobic surface patches, dynamic protein secondary structure monitoring, and the proclivity for solvent exposure of the proposed β-amyloid (Aβ) binding region of apolipoprotein A-I (apoA-I), “LN.” This study reveals that previously characterized ellipsoidal HDL3a and HDL2a models revert to a more spherical geometry in an atomistic representation due to the enhanced conformational flexibility afforded to the apoA-I protein secondary structure, allowing for enhanced surface lipid packing and lower overall surface hydrophobicity. Indeed, the proportional surface hydrophobicity and apoA-I exposure reduced with increasing HDL size, consistent with previous characterizations. Furthermore, solvent exposure of the “LN” region of apoA-I was exclusively limited to the smallest HDL3c model within the timescale of the simulations, and typically corresponded to a distinct loss in secondary structure across the “LN” region to form part of a significant contiguous hydrophobic patch on the HDL surface. Taken together, these findings provide preliminary evidence for a subpopulation-specific interaction between HDL3c particles and circulating hydrophobic species such as Aβ via the exposed “LN” region of apoA-I.

Covalent conjugate of pea protein induced by cyanidin-3-O-glucoside quinone: The structural formation and functional properties

The unique structural of anthocyanins makes them vulnerable to degradation within food processing environments. Proteins commonly coexist with polyphenols in natural food matrices on a non-covalent and covalent basis, serving to stabilize protection for polyphenols. Here, three natural pea-extracted proteins, including albumin, vicilin, and legumin were bound to cyanidin-3-O-glucoside (C3G), and binding modes, conjugate structures and functions were characterized. Isothermal titration calorimetry showed a strongly exothermic interaction between C3G and proteins. The results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fourier transform infrared spectroscopy revealed that the C3G-quinone (C3GQ) formed by C3G oxidation established covalent cross-links with C–S and C–N in the amino acid residues of proteins, and could form more large molecular weight aggregates at an addition amount of 1 mM. The free sulfhydryl and amino contents of the three protein conjugates decreased with the addition of C3GQ. Due to the presence of 4–5 times more sulfhydryl groups in albumin as compared to globulin, albumin showed a 20% higher binding capacity for C3GQ than globulin. This consequently led to the highest antioxidant activity in the C3GQ-albumin conjugate. In contrast to native pea proteins, the introduction of covalent modifications induces changes in the secondary structure of the pea proteins. Specifically, the structure of albumin tends to exhibit increased orderliness, while the molecular flexibility of globular proteins was augmented. Thus, the conjugates have enhanced thermal stability. Our results affirmed that the stability of C3G can be more effectively improved through designing it to covalently interact with proteins relative to non-covalent interactions. This study aimed at offering novel theoretical underpinnings for the functional utilization of C3G and pea proteins in the food industry.

Effect of thermal and defatting treatments on the composition, protein profile and structure of house cricket (Acheta domesticus) protein extracts

This study investigated the impact of blanching (100 °C, 40 s), defatting method (maceration, Soxhlet) and solvent polarity (hexane, ethanol) on the profile, structure and solubility of house cricket protein extracts. Blanching and Soxhlet using ethanol impacted the protein profile, with a lower content of myosin heavy chain and a higher abundance of low molecular weight proteins (<25 kDa). Moreover, ethanol induced aggregation of non-blanched cricket proteins, with a 13–72% reduction in protein recovery yield. The protein secondary structure of non-blanched extracts was also affected by ethanol with 18% more β-sheets. Furthermore, blanching resulted in a lower protein surface hydrophobicity by a factor of 3 to 7, with no impact of solvent polarity. Finally, the solubility of protein extracts remained >75%, regardless of the blanching and defatting methods. These findings, combined with the evaluation of techno-functional properties, could be used for the development of cricket-based protein ingredients for food formulations.

Effect of salting and dehydration treatments on the physicochemical and gel properties of hen and duck egg yolks, plasma and granules.

Salted hen egg yolks are less oily and less flavorful than salted duck egg yolks. However, hen eggs have a more adequate market supply and have a broader application prospect than duck eggs. In the present study, egg yolks, plasma, and granules were dehydrated by adding 1% NaCl to simulate traditional curing process of salted egg yolk. The changes in the pickling process of hen egg yolks (HEY) and duck egg yolks (DEY) plasma and granules were compared to reveal the gelation mechanism and the underlying causes of quality differences in salted HEY and DEY. Salted HEY can be compared with the changes in DEY during the pickling process to provide a theoretical basis for the quality improvement of salted HEY to salted DEY. The results showed that both plasma and granules were involved in gel formation, but exhibited different aggregation behaviors. Based on the intermolecular forces, the HEY proteins achieved aggregation mainly through hydrophobic interactions and DEY proteins mainly through covalent binding. According to spin-spin relaxation time, HEY gels immobilized a large amount of lipid and interacted strongly with lipids. DEY gels showed much free lipid and had weak interaction with lipid. The microstructure showed that HEY proteins were easily unfolded to form a homogeneous three-dimensional gel network structure after salting, whereas heterogeneous aggregates were formed to hinder the gel development in DEY. Changes in protein secondary structure content showed that pickling can promote the transformation of the α-helices to β-sheets structure in HEY gels, whereas more α-helices structure was formed in DEY gels. The present study has demonstrated that different gelation behaviors of hen and duck egg yolk proteins (especially in plasma) through salting treatment led to the difference in the quality of salted HEY and DEY. © 2024 Society of Chemical Industry.

The investigation of the molecular changes during lipopolysaccharide-induced systemic inflammation on rat hippocampus by using FTIR spectroscopy.

The aim of this study is to reveal the molecular changes accompanying the neuronal hyper-excitability during lipopolysaccharide (LPS)-induced systemic inflammation on rat hippocampus using Fourier transform infrared (FTIR) spectroscopy. For this aim, the body temperature of Wistar albino rats administered LPS or saline was recorded by radiotelemetry. The animals were decapitated when their body temperature began to decrease by 0.5°C after LPS treatment and the hippocampi of them were examined by FTIR spectroscopy. The results indicated that systemic inflammation caused lipid peroxidation, an increase in the amounts of lipids, proteins and nucleic acids, a decrease in membrane order, an increase in membrane dynamics and changes in the secondary structure of proteins. Principal component analysis successfully separated control and LPS-treated groups. In conclusion, significant structural, compositional and functional alterations occur in the hippocampus during systemic inflammation and these changes may have specific characteristics which can lead to neuronal hyper-excitability.

Evaluation of the Quality Changes in Three Commercial Pastourma Samples during Refrigerated Storage Using Physicochemical, Microbiological, and Image Analyses Combined with Chemometrics.

Despite the inherent stability of dried and cured products, such as pastourma, appropriate refrigeration remains essential for preserving their optimal characteristics. This study explored quality and safety characteristics in lamb, beef, and buffalo pastourma during 16-day refrigeration storage after package opening. The comprehensive approach employed Attenuated Total Reflection-Fourier-Transform Infrared (ATR-FTIR) spectroscopy, colorimetry, and image analysis, alongside physicochemical and microbiological analyses, to shed light on these alterations. The findings reveal a reduction in textural uniformity and color vibrancy (fading reds and yellows) across all samples during storage, with lamb pastourma exhibiting the most pronounced effects. Notably, image analysis emerged as a powerful tool, enabling the accurate classification of samples based on storage duration. Additionally, significant variations were observed in moisture content, hue angle, firmness, and TBARS levels, highlighting their influence on pastourma quality. The study documented a gradual decrease in lactic acid bacteria and aerobic plate count populations over time. ATR-FTIR spectra's interpretation revealed the presence of lipids, proteins, carbohydrates, and water. Protein secondary structures, demonstrably influenced by the meat type used, exhibited significant changes during storage, potentially impacting the functional and textural properties of pastourma. Overall, the findings contribute to a deeper understanding of pastourma spoilage during storage, paving the way for the development of improved preservation and storage strategies.

Rapid quantitative analysis of soybean protein isolates secondary structure by two-dimensional correlation infrared spectroscopy through pH perturbation

The infrared spectroscopy (IR) signal of protein is prone to being covered by impurity signals, and the accuracy of the secondary structure content calculated using spectral data is poor. To tackle this challenge, a rapid high-precision quantitative model for protein secondary structure was proposed. Firstly, a two-dimensional correlation calculation was performed based on 60 groups of soybean protein isolates (SPI) infrared spectroscopy data, resulting in a two-dimensional correlation infrared spectroscopy (2DCOS-IR). Subsequently, the optimal characteristic bands of the four secondary structures were extracted from the 2DCOS-IR. Ultimately, partial least squares (PLS), long short-term memory (LSTM), and bidirectional long short-term memory (BILSTM) algorithms were used to model the extracted characteristic bands and predict the content of SPI secondary structure. The findings suggested that BILSTM combined with 2DCOS-IR model (2DCOS-BILSTM) exhibited superior predictive performance. The prediction sets for α-helix, β-sheet, β-turn, and random coil were designated as 0.9257, 0.9077, 0.9476, and 0.8443, respectively, and their corresponding RMSEP values were 0.26, 0.48, 0.20, and 0.15. This strategy enhances the precision of IR and facilitates the rapid identification of secondary structure components within SPI, which is vital for the advancement of protein industrial production.

Cryoprotective effect of tamarind seed polysaccharide on grass carp surimi: characteristics, interactions, and mechanisms

In surimi products, the quality deterioration caused by freezing is a common problem. Therefore, the development of natural cryoprotectants is receiving increased attention. Tamarind seed polysaccharide (TSP) has been found to have a superior ice recrystallization inhibitory effect. In this work, the cryoprotective effect of TSP on grass carp surimi was investigated, including freeze-thaw stability, water distribution, texture, and microstructure. Additionally, the interaction between TSP and myofibrillar protein was analyzed. The inclusion of TSP resulted in the formation of a more regular and compact gel network through hydrogen bonding, which enhanced the binding between water molecules and myofibrillar protein. This, in turn, immobilized free water within the gel network. Furthermore, TSP prevented the texture deterioration of the surimi gel during repeated freeze-thaw cycles, particularly when 1.0% TSP was added. Moreover, the presence of TSP delayed the decline of Ca2+-ATPase activity and stabilized the secondary, tertiary, and quaternary structure of myofibrillar protein during freeze-thaw cycles. Therefore, TSP shows promise as a potential cryoprotectant for frozen surimi products, enhancing their quality and stability during frozen storage.

Unlocking the potential of waxworm (Galleria mellonella) proteins: Extraction, fractionation, and protein quality assessment

Edible insects are increasingly recognized as an alternative and sustainable protein source to meet future protein demands. Owing to its significant protein content (33.43%), the edible insect Waxworm (Galleria mellonella) was characterized in this study for its protein extractability, structural properties, and protein quality. Protein extractability under various pH levels and ionic strengths was evaluated. Subsequently, Osborne fractionation was employed to characterize protein fractions based on their preferential solubilities. The extracted proteins and their respective fractions were visualized by SDS-PAGE, and the most intense bands were excised for proteomics data analysis. Secondary structure, pI, molecular mass, and denaturation temperature were also examined. The quality of waxworm protein was assessed by determining the IVPD, PDCAAS, and amino acid profile. Waxworm proteins exhibited greater stability in alkaline conditions compared to acidic environments. These might also be more sensitive to ionic strength than other alternative proteins, such as those derived from plants. The allergen tropomyosin was identified by SDS-PAGE with an approximate molecular weight of ∼32.5 kDa; however, it can be selectively removed by salt extraction. The secondary structures of waxworm proteins, extracted under various conditions, appeared stable, with α-helix and β-conformation being the predominant conformation. Among the protein fractions, prolamins displayed an α-helix conformation, while albumins, globulins, and glutelins predominantly adopted a β-sheet structure. It was observed that structural features could influence the overall quality of the protein. As for the protein quality, globulins are the protein fraction recommended as these showed the highest EAAI% (924.02), PDCAAS (0.84) and IVPD (76.33%) values.

How does ionizing radiation affect amyloidogenesis in plants?

Purpose Ionizing radiation is a harsh environmental factor that could induce plant senescence. We hypothesized that radiation-related senescence remodels proteome, particularly by triggering the accumulation of prion-like proteins in plant tissues. The object of this study, pea (Pisum sativum L.), is an agriculturally important legume. Research on the functional importance of amyloidogenic proteins was never performed on this species. Materials and methods Pea seeds were irradiated in the dose range 5–50 Gy of X-rays. Afterward, Fourier-transform infrared spectroscopy (FTIR) was used to investigate changes in the secondary structure of proteins in germinated 3-day-old seedlings. Specifically, we evaluated the ratio between the amide I and II peaks. Next, we performed protein staining with Congo red to compare the presence of amyloids in the samples. In parallel, we profiled the detergent-resistant proteome fraction by ultrahigh-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS). Differentially accumulated proteins were functionally analyzed in MapMan software, and the PLAAC tool was used to predict putative prion-like proteins. Results We showed a reduced germination rate but higher plant height and faster appearance of reproductive organs in the irradiated at dose of 50 Gy group compared with the control; furthermore, we demonstrated more β-sheets and amyloid aggregates in the roots of stressed plants. We detected 531 proteins in detergent-resistant fraction extracted from roots, and 45 were annotated as putative prion-like proteins. Notably, 29 proteins were significantly differentially abundant between the irradiated and the control groups. These proteins belong to several functional categories: amino acid metabolism, carbohydrate metabolism, cytoskeleton organization, regulatory processes, protein biosynthesis, and RNA processing. Thus, the discovery proteomics provided deep data on novel aspects of plant stress biology. Conclusion Our data hinted that protein accumulation stimulated seedlings’ growth as well as accelerated ontogenesis and, eventually, senescence, primarily through translation and RNA processing. The increased abundance of primary metabolism-related proteins indicates more intensive metabolic processes triggered in germinating pea seeds upon X-ray exposure. The functional role of detected putative amyloidogenic proteins should be validated in overexpression or knockout follow-up studies.

SERT-StructNet: Protein secondary structure prediction method based on multi-factor hybrid deep model

Protein secondary structure prediction (PSSP) is a pivotal research endeavour that plays a crucial role in the comprehensive elucidation of protein functions and properties. Current prediction methodologies are focused on deep-learning techniques, particularly focusing on multi-factor features. Diverging from existing approaches, in this study, we placed special emphasis on the effects of amino acid properties and protein secondary structure propensity scores (SSPs) on secondary structure during the meticulous selection of multi-factor features. This differential feature-selection strategy results in a distinctive and effective amalgamation of the sequence and property features. To harness these multi-factor features optimally, we introduced a hybrid deep feature extraction model. The model initially employs mechanisms such as dilated convolution (D-Conv) and a channel attention network (SENet) for local feature extraction and targeted channel enhancement. Subsequently, a combination of recurrent neural network variants (BiGRU and BiLSTM), along with a transformer module, was employed to achieve global bidirectional information consideration and feature enhancement. This approach to multi-factor feature input and multi-level feature processing enabled a comprehensive exploration of intricate associations among amino acid residues in protein sequences, yielding a Q3 accuracy of 84.9% and an Sov score of 85.1%. The overall performance surpasses that of the comparable methods. This study introduces a novel and efficient method for determining the PSSP domain, which is poised to deepen our understanding of the practical applications of protein molecular structures.

Effects of acidification by traditional Jiaozi starter and neutralization with alkali (Na2CO3) on whole wheat dough properties.

Whole wheat steamed bread has been recommended for its potential nutritional benefits to human health. Given the positive role of both organic acid and alkali in improving dough development and product quality, the present study investigated the effects of neutralization by addition of alkali (Na2CO3) after dough acidification with traditional Jiaozi starter on the properties of whole wheat dough. The population of yeast and lactic acid bacteria and the acidification level of the dough increased significantly after fermentation with Jiaozi. Incorporation of alkali greatly improved the leavening capacity of the remixed dough and the quality of steamed bread. Jiaozi fermentation and alkali addition changed the water distribution patterns (T2) and affected the secondary structures of gluten protein, starch crystallinity and pasting properties. The storage modulus (G') of the dough increased significantly with the alkali addition, which could be attributed to the promoted cross-linking of the gluten structure and the altered hydration state of the macromolecules. The results of the present study indicate that a combination of Jiaozi fermentation and alkali addition could improve the technological properties of whole wheat dough and the quality of steamed bread. The results will help us to further explore the potential application of moderate acidification and alkali addition in the production of leavened whole wheat products. © 2024 Society of Chemical Industry.

Freezing-mediated formation of supraproteins using depletion forces

HypothesisLong-acting formulations such as microparticles, injectable depots and implantable devices can realize spatiotemporally controlled delivery of protein drugs to extend their therapeutic in vivo half-lives. To efficiently encapsulate the protein drugs into such drug delivery systems, (sub)micron-sized protein particles are needed. The formation of micronized supraproteins can be induced through the synergistic combination of attractive depletion forces and freezing. The size of the supraproteins can be fine-tuned from submicron to several microns by adjusting the ice crystallization rate through the freeze-quench depth, which is set by the target temperature.MethodsSupraprotein micron structures were prepared from protein solutions under various conditions in the presence and absence of nonadsorbing polyethylene glycol. Scanning electron microscopy and dynamic light scattering were employed to determine the sizes of the supraproteins and real-time total internal reflection fluorescent microscopy was used to follow the supraprotein formation during freezing. The protein secondary structure was measured before and after micronization by circular dichroism. A phase diagram of a protein–polyethylene glycol mixture was theoretically predicted to investigate whether the depletion interaction can elucidate the phase behavior.FindingsMicronized protein supraparticles could be prepared in a controlled manner by rapid freeze-drying of aqueous mixtures of bovine serum albumin, horseradish peroxidase and lysozyme mixed with polyethylene glycol. Upon freezing, the temperature quench initiates a phase separation process which is reminiscent of spinodal decomposition. This demixing is subsequently arrested during droplet phase separation to form protein-rich microstructures. The final size of the generated protein microparticles is determined by a competition between phase separation and cooling rate, which can be controlled by target temperature. The experimental phase diagram of the aqueous protein–polyethylene glycol dispersion aligns with predictions from depletion theory for charged colloids and nonadsorbing polymers.

Effects of high-pressure homogenization and ultrasound on the composition, structure, and physicochemical properties of proteins extracted from Nannochloropsis Oceania

This study examined the effects of high-pressure homogenization (HPH) and ultrasonication pre-treatment on the structural and physicochemical properties of proteins extracted from defatted Nannochloropsis Oceania biomass (DNOB). HPH treatment was found to enhance the solubility of protein extracted from DNOB compared to ultrasound, where samples pretreated with three passes (3P) of HPH exhibited lower solubility than two passes (2P). The morphology of extracted samples was visualized by scanning electron microscopy, which HPH pre-treatment, especially with more passes, were able to breakdown DNOB into fragments. Alternatively, more holes were displayed on the surface of the extracts pretreated with ultrasound especially when higher amplitude applied. The particle size of extracts from HPH3P (129.5 µm) significant dropped from HPH2P (314.25 µm), where samples pretreated with ultrasound at 20 % amplitude (US20) also decreased in particle size compared to 40 % amplitude (US40), from 115.25 µm to 78.22 µm. Protein flexibility of DNOB extracts were enhanced by both HPH2P and HPH3P but decreased for ultrasound samples. β-sheets were found to be the most abundant protein secondary structure for all samples, where samples treated with HPH3P contained the highest percentage of β-sheets (72 %) than control, HPH2P, ultrasonication at 20 and 40 % amplitude (52–62 %). The high percentage of β-sheets found in HPH3P sample also contributed to its outstanding emulsifying properties which stood out among all, especially at concentrations over 1 mg/ml. Results obtained from this study helped to direct the application of DNOB extracts as functional food ingredient for future food innovation.

PhosAF: An integrated deep learning architecture for predicting protein phosphorylation sites with AlphaFold2 predicted structures

Phosphorylation is indispensable in comprehending biological processes, while biological experimental methods for identifying phosphorylation sites are tedious and arduous. With the rapid growth of biotechnology, deep learning methods have made significant progress in site prediction tasks. Nevertheless, most existing predictors only consider protein sequence information, that limits the capture of protein spatial information. Building upon the latest advancement in protein structure prediction by AlphaFold2, a novel integrated deep learning architecture PhosAF is developed to predict phosphorylation sites in human proteins by integrating CMA-Net and MFC-Net, which considers sequence and structure information predicted by AlphaFold2. Here, CMA-Net module is composed of multiple convolutional neural network layers and multi-head attention is appended to obtaining the local and long-term dependencies of sequence features. Meanwhile, the MFC-Net module composed of deep neural network layers is used to capture the complex representations of evolutionary and structure features. Furthermore, different features are combined to predict the final phosphorylation sites. In addition, we put forward a new strategy to construct reliable negative samples via protein secondary structures. Experimental results on independent test data and case study indicate that our model PhosAF surpasses the current most advanced methods in phosphorylation site prediction.