Stable Interstrand Cross-Links Generated from the Repair of 1,N6-Ethenoadenine in DNA by α-Ketoglutarate/Fe(II)-Dependent Dioxygenase ALKBH2.

DNA cross-links severely challenge replication and transcription in cells, promoting senescence and cell death. In this paper, we report a novel type of DNA interstrand cross-link (ICL) produced as a side product during the attempted repair of 1,N6-ethenoadenine (εA) by human α-ketoglutarate/Fe(II)-dependent enzyme ALKBH2. This stable/nonreversible ICL was characterized by denaturing polyacrylamide gel electrophoresis analysis and quantified by high-resolution LC-MS in well-matched and mismatched DNA duplexes, yielding 5.7% as the highest level for cross-link formation. The binary lesion is proposed to be generated through covalent bond formation between the epoxide intermediate of εA repair and the exocyclic N6-amino group of adenine or the N4-amino group of cytosine residues in the complementary strand under physiological conditions. The cross-links occur in diverse sequence contexts, and molecular dynamics simulations rationalize the context specificity of cross-link formation. In addition, the cross-link generated from attempted εA repair was detected in cells by highly sensitive LC-MS techniques, giving biological relevance to the cross-link adducts. Overall, a combination of biochemical, computational, and mass spectrometric methods was used to discover and characterize this new type of stable cross-link both in vitro and in human cells, thereby uniquely demonstrating the existence of a potentially harmful ICL during DNA repair by human ALKBH2.

Interactions among the composition changes in fungal communities and the main mycotoxins in simulated stored wheat grains.

BACKGROUND: There are significant food safety risks associated with wheat spoilage due to fungal growth and mycotoxin contamination. Nevertheless, a few studies have examined how stored wheat grain microbial communities and mycotoxins vary in different storage conditions. In this study, changes in deoxynivalenol (DON) and deoxynivalenol-3-glucoside (D3G) content were measured with ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS), and an amplicon sequence analysis of fungi was performed on stored wheat grains from different storage conditions using high-throughput sequencing. The detailed interactions among the composition changes in the fungal community and the DON content of simulated stored wheat grains were also analyzed. RESULTS: Alternaria, Fusarium, Mrakia, and Aspergillus were the core fungal taxa, and the fungal communities of samples stored under different conditions were observed to be different. Aspergillus relative abundances increased, whereas Fusarium decreased. This led to an increase in the content of DON. The content of DON increased about 67% with 12% moisture and at 25 °C after 2 months of storage, which was influenced by the stress response of Fusarium. Correlations in fungal and mycotoxins changes were observed. There may be potential value in these findings for developing control strategies to prevent mildew infestations and mycotoxins contamination during grain storage. CONCLUSION: In storage, the more the fungal community composition and the relative abundance of Fusarium change, the more mycotoxins will be produced. We should therefore reduce competition between fungal communities through pre-storage treatment and through measures during storage. © 2023 Society of Chemical Industry.

REV7 Monomer Is Unable to Participate in Double Strand Break Repair and Translesion Synthesis but Suppresses Mitotic Errors.

Rev7 is a regulatory protein with roles in translesion synthesis (TLS), double strand break (DSB) repair, replication fork protection, and cell cycle regulation. Rev7 forms a homodimer in vitro using its HORMA (Hop, Rev7, Mad2) domain; however, the functional importance of Rev7 dimerization has been incompletely understood. We analyzed the functional properties of cells expressing either wild-type mouse Rev7 or Rev7K44A/R124A/A135D, a mutant that cannot dimerize. The expression of wild-type Rev7, but not the mutant, rescued the sensitivity of Rev7-/- cells to X-rays and several alkylating agents and reversed the olaparib resistance phenotype of Rev7-/- cells. Using a novel fluorescent host-cell reactivation assay, we found that Rev7K44A/R124A/A135D is unable to promote gap-filling TLS opposite an abasic site analog. The Rev7 dimerization interface is also required for shieldin function, as both Rev7-/- cells and Rev7-/- cells expressing Rev7K44A/R124A/A135D exhibit decreased proficiency in rejoining some types of double strand breaks, as well as increased homologous recombination. Interestingly, Rev7K44A/R124A/A135D retains some function in cell cycle regulation, as it maintains an interaction with Ras-related nuclear protein (Ran) and partially rescues the formation of micronuclei. The mutant Rev7 also rescues the G2/M accumulation observed in Rev7-/- cells but does not affect progression through mitosis following nocodazole release. We conclude that while Rev7 dimerization is required for its roles in TLS, DSB repair, and regulation of the anaphase promoting complex, dimerization is at least partially dispensable for promoting mitotic spindle assembly through its interaction with Ran.

Modification of Soft Wheat Protein for Improving Cake Quality by Superheated Steam Treatment of Wheat Grain.

Many varieties of soft wheat in China cannot fully satisfy the requirements of making high-quality cakes due to their undesirable protein properties, which leads to shortages of high-quality soft wheat flour. Therefore, a modification of soft wheat protein is essential for improving the quality of soft wheat and thus improving cake quality. In order to modify the protein properties of soft wheat used for cake production, superheated steam (SS) was used to treat soft wheat grains at 165 °C and 190 °C for 1, 2, 3, 4, and 5 min, respectively, followed by the milling of wheat grains to obtain refined wheat flour. The properties of proteins and cakes were analyzed using refined wheat flour as materials. First, changes in the structures of wheat proteins were analyzed by determining the solubility, molecular weight distribution and secondary structure of proteins in wheat flour. Secondly, changes in the functional properties of proteins were analyzed by determining the foaming properties and emulsifying properties of proteins in wheat flour. Finally, the specific volume and texture of cakes with wheat flour milled from SS-treated wheat were analyzed. At the initial stage of SS treatment, some of the gliadin and glutenin aggregated, and the gluten macro-polymer (GMP) contents increased. This allowed a more stable gluten network to form during dough kneading, leading to an improvement in dough elasticity. In addition, a short time period (1-3 min) of SS treatment improved the emulsifying properties and foaming ability of wheat protein, which helped to improve the specific volume and texture of cakes. Increasing the SS temperature from 165 °C to 190 °C reduced the optimal treatment time needed to improve cake quality from 3 min to 1 min. SS treatment for longer time (>3 min) periods led to severe protein aggregation and a decrease in the foaming ability and emulsifying properties of protein, which led to a deterioration in the cake quality. Thus, SS treatment at 165 °C for 1-3 min and 190 °C for 1 min could be a suitable method of improving the physicochemical properties of soft wheat used to make cakes with high specific volumes and good texture.

Structure-activity relationship between gluten and dough quality of sprouted wheat flour based on air classification-induced component recombination.

BACKGROUND: Air classification can separate sprouted wheat flour (SWF) into three types: coarse wheat flour (F1), medium wheat flour (F2) and fine wheat flour (F3). The gluten quality of SWF can be indirectly improved by removing inferior parts (F3). In order to reveal the underlying mechanism of this phenomenon, the composition and structural changes of gluten, as well as the rheological properties and fermentation characteristics of gluten in recombinant dough in the process of air classification of all three SWF types, were analyzed in this study. RESULTS: Overall, sprouting significantly reduced the content of high-molecular-weight subunits, such as glutenin subunit and ω-gliadin. It also destroyed the structural content, such as disulfide bonds, α-helix and ß-turn contents, which maintained the stability of gluten gel. Air classification made the above changes in F3 more severe but reversed them in F1. Moreover, rheological properties were more affected by gluten composition, whereas fermentation characteristics were more affected by gluten structure. CONCLUSION: After air classification, particles rich in high molecular weight subunits from SWF are enriched in F1, and the gluten of F1 has more secondary structure that maintain gel stability, which ultimately lead to improved rheology properties and fermentation characteristics. F3 relatively exhibits the opppsite phenomenon. These results further reveal the potential mechanism of improvement of SWF gluten by air classification. Moreover, Thus, this study provides new perspectives for the utilization of SWF. © 2023 Society of Chemical Industry.

Superheated steam processing of cereals and cereal products: A review.

The concept of superheated steam (SS) was proposed over a century ago and has been widely studied as a drying method. SS processing of cereals and cereal products has been extensively studied in recent years for its advantages of higher drying rates above the inversion temperature, oxygen-free environment, energy conservation, and environmental protection. This review provides a brief introduction to the history, principles, and classification of SS. The applications of SS processing in the drying, enzymatic inactivation, sterilization, mycotoxin degradation, roasting, and cooking of cereals and cereal products are summarized and discussed. Moreover, the effects of SS processing on the physicochemical properties of cereals and the qualities of cereal foods are reviewed and discussed. The applications of SS for cereal processing and its effects on cereal properties have been extensively studied; however, issues such as the browning of cereal foods, thermal damage of starch, protein denaturation, and nutrition loss have not been comprehensively studied. Therefore, further studies are required to better understand the mechanism of the quality changes caused by SS processing and to expand the fields of application of SS in the cereal processing industry. This review enhances the understanding of SS processing and presents theoretical suggestions for promoting SS processing to improve the safety and quality of cereals and cereal products.

Behavior of wheat flour dough at different pretreated temperatures through rheological characteristics and molecular interactions of proteins.

Rheological properties and chemical interactions of doughs prepared at different temperatures were evaluated. The results showed that the rigidity of pretreated doughs was enhanced, and the processing performance of doughs was weakened. Preheating resulted in the polymerization of gluten through the conversion of sulfhydryl groups to disulfide bonds. The noncovalent interaction of dough played a dominant role and further led to the production of glutenin macropolymers (55.77 mg/g). CLSM images verified that preheating promoted the formation of the coarse and scattered gluten network, while preheating at 80 °C led to a higher gluten area percentage (40.27 %) and lower lacunarity (6.74 × 10-2) structure. The migration of water promoted changes in hydrogen bond and hydrophobic interaction in doughs, which directly affect the processability of doughs. The study provides information for predicting the rheological behavior of dough in actual production and makes it possible to modify gluten by preheating treatment without complicating existing operations.

Reduction of Aflatoxin B1 and Zearalenone Contents in Corn Using Power Ultrasound and Its Effects on Corn Quality.

The degradation of aflatoxin B1 (AFB1) and zearalenone (ZEA) is investigated using power ultrasound to identify suitable methods to reduce the mycotoxin content of corn. AFB1 and ZEA in corn are simultaneously degraded via power ultrasound; thus, this method has a significant effect on corn quality. The power intensity, solid-liquid ratio, and ultrasonic treatment modes significantly affect the degradation rates of AFB1 and ZEA. The dissolution of AFB1 and ZEA in water also facilitates their degradation. At the initial stage of ultrasonic treatment, power ultrasound promotes the dissolution of mycotoxins in water, whereupon they are partially oxidized by free radicals. With a treatment time of 10 min, the reduction rates decreased owing to the dissolution of combined-state mycotoxins. After ultrasonic treatment, the contents of the essential amino acids, the total number of amino acids, and the fatty acids in corn decreased; however, ΔH values decreased during starch gelatinization. In contrast, the amylose content and viscosity of corn significantly increased during gelatinization. Therefore, this method is potentially suitable for the reduction of AFB1 and ZEA contents in corn.

A haem-sequestering plant peptide promotes iron uptake in symbiotic bacteria.

Symbiotic partnerships with rhizobial bacteria enable legumes to grow without nitrogen fertilizer because rhizobia convert atmospheric nitrogen gas into ammonia via nitrogenase. After Sinorhizobium meliloti penetrate the root nodules that they have elicited in Medicago truncatula, the plant produces a family of about 700 nodule cysteine-rich (NCR) peptides that guide the differentiation of endocytosed bacteria into nitrogen-fixing bacteroids. The sequences of the NCR peptides are related to the defensin class of antimicrobial peptides, but have been adapted to play symbiotic roles. Using a variety of spectroscopic, biophysical and biochemical techniques, we show here that the most extensively characterized NCR peptide, 24 amino acid NCR247, binds haem with nanomolar affinity. Bound haem molecules and their iron are initially made biologically inaccessible through the formation of hexamers (6 haem/6 NCR247) and then higher-order complexes. We present evidence that NCR247 is crucial for effective nitrogen-fixing symbiosis. We propose that by sequestering haem and its bound iron, NCR247 creates a physiological state of haem deprivation. This in turn induces an iron-starvation response in rhizobia that results in iron import, which itself is required for nitrogenase activity. Using the same methods as for L-NCR247, we show that the D-enantiomer of NCR247 can bind and sequester haem in an equivalent manner. The special abilities of NCR247 and its D-enantiomer to sequester haem suggest a broad range of potential applications related to human health.

Comparison and mechanism analysis of the changes in viscoelasticity and texture of fresh noodles induced by wheat flour lipids.

To understand the functionality of wheat flour starch lipids (SLs), non-starch lipids (NSLs), glycolipids (GLs), phospholipids (PLs), and neutral lipids (NLs) in non-leavened wheat-based products, their independent influence on noodle dough viscoelasticity and noodle texture were compared and the underlying mechanism was elucidated. Defatting caused slightly improved hydration, marginally promoting dough viscoelasticity and noodle springiness and adhesiveness but the resulting absence of starch-lipid complexes and few B-starch granules signally reduced the noodle hardness. Independently adding 2.50 g of these five lipids back into 100 g of defatted flour, GLs showed the most improved effects, followed by PLs, SLs, NSLs, and NLs. These lipids associated with gluten proteins and enhanced water-solids interplay, resulting in a significantly decreased SDS-soluble gluten proteins and further producing dough with increased ß-turn, moderate protein aggregation, and properly intensive microstructures. Consequently, the resultant noodle dough exhibited an optimal viscoelasticity and the cooked noodle had a desirable texture.

Mechanical action on the development of dough andits influence on rheological properties and protein network structure.

Four simple dough preparation methods were proposed to imitate the rheological behaviors of traditional hand-made doughs and the underlying mechanism was concomitantly elucidated. It indicated the hand-made doughs, including the conventional hand-made dough (CHD), bidirectional pressed dough (BPD), bidirectional rolled dough (BRD), unidirectional pressed dough (UPD), and unidirectional rolled dough (URD), showed weaker mechanical resistance than the mixer-made dough did. Compared with UPD and BRD, BPD and URD had better tensile resistance and deformation recovery. CLSM analysis showed that these two doughs also possessed smaller lacunarity (7.22-7.24 × 10-2) and larger branching rate (0.23 × 10-2), suggesting bidirectionally pressing and unidirectionally rolling could produce a dough with better gluten network connectivity. Analysis of gluten protein solubility showed that the stronger hydrogen bonds and hydrophobic interactions of gluten protein were derived in rolled doughs (URD and BRD), and the stronger slip caused by intermediate water in pressed doughs (UPD and BPD) may lead to the high gluten extractability. In addition, more disulfide bonds were formed in BPD (3.37 µmol/g) and URD (3.62 µmol/g), promoting the stronger mechanical resistance in BPD and URD. Nevertheless, pressing or rolling promoted no statistically significant increase in the content of glutenin macropolymers. Physical entanglement caused by the recombination of noncovalent interactions may be the main cause. In conclusion, theeffect ofmanual external forces ondough qualitywasverified theoretically, and gluten network analysis can quantitatively evaluate dough microstructural changes.

Tinea of vellus hair: shining under the Wood's lamp.

We herein reported a simple and cheap method to diagnose the tinea of vellus hair, which is long-neglected and always wrongly treated.

A qPlus-based scanning probe microscope compatible with optical measurements.

We design and develop a scanning probe microscope (SPM) system based on the qPlus sensor for atomic-scale optical experiments. The microscope operates under ultrahigh vacuum and low temperature (6.2 K). In order to obtain high efficiency of light excitation and collection, two front lenses with high numerical apertures (N.A. = 0.38) driven by compact nano-positioners are directly integrated on the scanner head without degrading its mechanical and thermal stability. The electric noise floor of the background current is 5 fA/Hz1/2, and the maximum vibrational noise of the tip height is below 200 fm/Hz1/2. The drift of the tip-sample spacing is smaller than 0.1 pm/min. Such a rigid scanner head yields small background noise (oscillation amplitude of ∼2 pm without excitation) and high quality factor (Q factor up to 140 000) for the qPlus sensor. Atomic-resolution imaging and inelastic electron tunneling spectroscopy are obtained under the scanning tunneling microscope mode on the Au(111) surface. The hydrogen-bonding structure of two-dimensional (2D) ice on the Au(111) surface is clearly resolved under the atomic force microscope (AFM) mode with a CO-terminated tip. Finally, the electroluminescence spectrum from a plasmonic AFM tip is demonstrated, which paves the way for future photon-assisted SPM experiments.

Interactions between wheat globulin and gluten under alkali or salt condition and its effects on noodle dough rheology and end quality.

The influences of wheat globulin on dough and noodle quality under alkali or salt conditionwere investigated, and the protein interactions were revealed. Results indicated that dough viscoelasticity, noodle hardness, springiness and extensibility of samples with globulin added were remarkably increased under alkali condition. However, the corresponding enhancement was less significant under salt condition. In dough system, added globulin decreased the protein surface hydrophobicity by 38.71%, implying the enhancement of hydrophobic interactions. Under salt and alkali conditions, added globulin further increased the ß-sheets structure by 1.68% and 3.17%, respectively, indicating the enhancement of hydrogen bonds interaction. In addition, disulfide bonds interactions between globulin and gluten have also been demonstrated induced by alkali. The results were accountable for protein network polymerization observed in micro-structures. This paper provides new insights into the structural properties of wheat globulin, and demonstrates the excellent potential to improve noodle processing quality under alkali condition significantly.

Sequence Dependent Repair of 1,N6-Ethenoadenine by DNA Repair Enzymes ALKBH2, ALKBH3, and AlkB.

Mutation patterns of DNA adducts, such as mutational spectra and signatures, are useful tools for diagnostic and prognostic purposes. Mutational spectra of carcinogens derive from three sources: adduct formation, replication bypass, and repair. Here, we consider the repair aspect of 1,N6-ethenoadenine (εA) by the 2-oxoglutarate/Fe(II)-dependent AlkB family enzymes. Specifically, we investigated εA repair across 16 possible sequence contexts (5'/3' flanking base to εA varied as G/A/T/C). The results revealed that repair efficiency is altered according to sequence, enzyme, and strand context (ss- versus ds-DNA). The methods can be used to study other aspects of mutational spectra or other pathways of repair.

Effects of wheat flour particle size on flour physicochemical properties and steamed bread quality.

In this study, differently sized particles of wheat flour (from 52.36 µm to 108.89 µm) were obtained by adjusting the distance between the rolls (0.02, 0.04, 0.06, 0.08, and 0.1 mm) of a heart mill. Results showed that reducing the particle size significantly increased the damaged starch (DS) content. Uniaxial tensile measurement of dough showed that reducing the particle size of wheat flour can effectively increase the maximum tensile resistance, but the extensibility reaches the maximum in samples at medium particle diameter (78 and 66 µm). Additionally, the ratio of dynamic moduli (G″/G') decreased with a reducing particle size. The results of disulfide bond content, gluten microstructure, showed that finer flour granulation can strengthen the gluten network. The steamed bread (SB) making test showed that SB made from wheat flour of a smaller particle size had a significantly smaller specific volume than that made from a larger particle size. The texture profile analysis showed that with a decrease of wheat flour particle size, the hardness, chewiness of SB increased, the resilience decreased, and there was no significant difference in adhesiveness. Overall, the quality of SB made flour of medium particles (78 µm) is better.

Impact of wheat globulin addition on dough rheological properties and quality of cooked noodles.

Impact of globulin addition on the functional and protein structural properties of dough and cooked noodles were investigated. The underlying mechanism was explored through analyzing the interaction between globulin and gluten by using SDS-PAGE, size exclusion chromatography, free sulfhydryl/disulfide bond analysis, laser scanning confocal microscopy and Fourier transform infrared spectroscopy. Results showed that the stiffness/hardness and maximum resistance of dough and cooked noodles were both increased when globulin addition was 1.5% or higher. Besides, extensibility of cooked noodles was also improved when the addition up to 3.0%. The addition of globulin facilitated weakening the S-S bonds in the gluten network and cross-linked with SDS-soluble gluten mainly through non-covalent interactions, especially hydrophobic interactions. Meanwhile, a more rigid protein network structure was observed. Additionally, following cooking, globulin addition accelerated the aggregation of protein molecules. When the addition reached 3%, the protein conformation was transformed from ß-sheets and random coils to ß-turns.

Effects of vacuum degree, mixing speed, and water amount on the moisture distribution and rheological properties of wheat flour dough.

Effects of vacuum degrees (0.00, 0.02, 0.04, 0.06, 0.08 MPa) on water distribution state, tensile properties, stress relaxation properties, and viscoelasticity of dough, as well as the effects of mixing speed (50, 70, 90 rpm/min) and water content (40%, 45%, 50%) under optimum vacuum degree were studied. The results showed that the proper vacuum degree (0.06 MPa) could promote the full contact between flour and water and improved the water-holding capacity of the dough. Meanwhile, the dough had stronger tensile strength, the best viscoelasticity and the ability to recover from external deformation more quickly. Under the vacuum of 0.06 MPa, with the increasing of mixing speed, the response to the external force of dough increased first and then decreased. Adding more water reduced the strength of dough, weakened the response to external forces, and led to a significant decrease in tensile resistance and tensile area of the dough, as well as a decrease in viscoelasticity (p < 0.05). The proper vacuum mixing allowed the preparation of dough to require more water and less energy. PRACTICAL APPLICATION: In the processing of wheat flour products, vacuum mixing is considered to be beneficial to the quality of noodles and breads. As the intermediate of these products, the dough is of great significance for the monitoring of its rheological characteristics. In this study, a moderate vacuum degree led to a significant improvement in the rheological properties of the dough, and the processing performance was the best. Under the optimal vacuum degree, the influence of mixing speed and water amount cannot be ignored. Vacuum mixing is an efficient dough preparation method, which can produce certain economic benefits.

Interaction between A-type/B-type starch granules and gluten in dough during mixing.

To explore the interaction between A/B starch and gluten, the rheological and structural properties of starch-gluten dough with varied A/B starch ratios during mixing were investigated. The G' and G″ values of under- and overdeveloped dough with an A/B starch ratio of 5:5 were higher than those of dough with other ratios and decreased as the A/B starch ratio increased in optimized dough. B starch enhanced extension resistance and dough firmness. Small B starch granules promoted continuous gluten network formation, while large A starch granules readily separate from the gluten network. B starch promoted GMP polymerization. Covalent bonds were the main force involved in A starch-gluten interactions. Hydrophobic interactions were the main force in the under- to optimum-mixing stages, whereas hydrogen and covalent bonds were involved in B starch-gluten interactions from the optimum- to over-mixing stages. A model describing the interactions between gluten and starch components was proposed.

Nanoscale electric-field imaging based on a quantum sensor and its charge-state control under ambient condition.

Nitrogen-vacancy (NV) centers in diamond can be used as quantum sensors to image the magnetic field with nanoscale resolution. However, nanoscale electric-field mapping has not been achieved so far because of the relatively weak coupling strength between NV and electric field. Here, using individual shallow NVs, we quantitatively image electric field contours from a sharp tip of a qPlus-based atomic force microscope (AFM), and achieve a spatial resolution of ~10 nm. Through such local electric fields, we demonstrated electric control of NV's charge state with sub-5 nm precision. This work represents the first step towards nanoscale scanning electrometry based on a single quantum sensor and may open up the possibility of quantitatively mapping local charge, electric polarization, and dielectric response in a broad spectrum of functional materials at nanoscale.