Meta-QTL analysis and identification of candidate genes for multiple-traits associated with spot blotch resistance in bread wheat.

In bread wheat, a literature search gave 228 QTLs for six traits, including resistance against spot blotch and the following five other related traits: (i) stay green; (ii) flag leaf senescence; (iii) green leaf area duration; (iv) green leaf area of the main stem; and (v) black point resistance. These QTLs were used for metaQTL (MQTL) analysis. For this purpose, a consensus map with 72,788 markers was prepared; 69 of the above 228 QTLs, which were suitable for MQTL analysis, were projected on the consensus map. This exercise resulted in the identification of 16 meta-QTLs (MQTLs) located on 11 chromosomes, with the PVE ranging from 5.4% (MQTL7) to 21.8% (MQTL5), and the confidence intervals ranging from 1.5 to 20.7 cM (except five MQTLs with a range of 36.1-57.8 cM). The number of QTLs associated with individual MQTLs ranged from a maximum of 17 in MQTL3 to 8 each in MQTL5 and MQTL8 and 5 each in MQTL7 and MQTL14. The 16 MQTLs, included 12 multi-trait MQTLs; one of the MQTL also overlapped a genomic region carrying the major spot blotch resistance gene Sb1. Of the total 16 MQTLs, 12 MQTLs were also validated through marker-trait associations that were available from earlier genome-wide association studies. The genomic regions associated with MQTLs were also used for the identification of candidate genes (CGs) and led to the identification of 516 CGs encoding 508 proteins; 411 of these proteins are known to be associated with resistance against several biotic stresses. In silico expression analysis of CGs using transcriptome data allowed the identification of 71 differentially expressed CGs, which were examined for further possible studies. The findings of the present study should facilitate fine-mapping and cloning of genes, enabling Marker Assisted Selection.

Explaining the improving effect of dough crumb-sheet composite rolling on fresh noodle quality: From microstructure and moisture distribution perspective.

A new technique known as dough crumb-sheet composite rolling (DC-SCR) was used to improve the quality of fresh noodles. However, there is a dearth of theoretical investigations into the optimal selection of specific parameters for this technology, and the underlying mechanisms are not fully understood. Therefore, the effects of dough crumb addition times in DC-SCR on the texture, cooking, and eating quality of fresh noodles were first studied. Then, the underlying regulation mechanism of DC-SCR technology on fresh noodles was analyzed in terms of moisture distribution and microstructure. The study demonstrated that the most significant enhancement in the quality of fresh noodles was achieved by adding dough crumbs six times. Compared with fresh noodles made without the addition of dough crumbs, the initial hardness and chewiness of fresh noodles made by adding six times of dough crumbs increased by 25.32% and 46.82%, respectively. In contrast, the cooking time and cooking loss were reduced by 28.45% and 29.69%, respectively. This quality improvement in fresh noodles made by DC-SCR came from the microstructural differences of the gluten network between the inner and outer layers of the dough sheet. A dense structure on the outside and a loose structure on the inside could endow the fresh noodles made by DC-SCR with higher hardness, a shortened cooking time, and less cooking loss. This study would provide a theoretical and experimental basis for creating high-quality fresh noodles.

The impact of addition of pearl millet starch-germ complex in white bread on nutritional, textural, structural, and glycaemic response: Single blinded randomized controlled trial in healthy and pre-diabetic participants.

The rise of pre-diabetes at the global level has created a significant interest in developing low glycaemic index food products. The pearl millet is a cheaper source of starch and its germ contains significant amount of protein and fat. The complexing of pearl millet starch and germ by dry heat treatment (PMSGH) resulted an increase in the resistant starch content upto 45.09 % due to formation of amylose-glutelin-linoleic acid complex. The resulting pearl millet starch germ complex was incorporated into wheat bread at 20, 25, and 30 %. The PMSGH incorporated into bread at 30 % reduced the glycaemic index to 52.31. The PMSGH incorporated bread had significantly (p < 0.05)increased in the hardness with a reduction in springiness and cohesiveness. The structural attributes of the 30 % PMSGH incorporated bread revealed a significant (p < 0.05)increase in 1040/1020 cm-1 ratio and relative crystallinity. The consumption of functional bread incorporated with pearl millet starch germ complex reduced blood glucose levels and in vivo glycaemic index in healthy and pre-diabetic participants when compared to white bread. Hence, the study showed that the incorporation of pearl millet starch-germ complex into food products could be a potential new and healthier approach for improving dietary options in pre-diabetes care.

Heat-moisture treatment can modulate all-purpose wheat flour for short dough biscuit making: Evidences and mechanism.

In view of the merits of all-purpose wheat flour (APWF) to soft wheat flour (SWF) in cost and protein supply, the feasibility of heat-moisture treatment (HMT, 19% moisture for 1 h at 60, 80 and 100 °C, respectively) to modify APWF as a substitute SWF in making short dough biscuits was explored. For underlying mechanisms, on the one hand, HMT reduced the hydration capacity of damaged starch particles by coating them with denatured proteins. On the other hand, HMT at 80 °C and 100 °C significantly denatured gluten proteins to form protein aggregates, highly weakening the gluten network in dough. These two aspects jointly conferred APWF dough with higher deformability and therefore significantly improved the qualities of biscuits. Moreover, the qualities of biscuits from APWF upon HMT-100 °C were largely comparable to that from SWF, even higher values were concluded in spread ratio, volume, specific volume and consumer acceptance.

Deciphering the inhibitory mechanisms of betanin and phyllocactin from Hylocereus polyrhizus peel on protein glycation, with insights into their application in bread.

Protein glycation closely intertwines with the pathogenesis of various diseases, sparking a growing interest in exploring natural antiglycation agents. Herein, high-purity betacyanins (betanin and phyllocactin) derived from Hylocereus polyrhizus peel were studied for their antiglycation potential using an in vitro bovine serum albumin (BSA)-glucose model. Notably, betacyanins outperformed aminoguanidine, a recognized antiglycation agent, in inhibiting glycation product formation across different stages, especially advanced glycation end-products (AGEs). Interestingly, phyllocactin displayed stronger antiglycation activity than betanin. Subsequent mechanistic studies employing molecular docking analysis and fluorescence quenching assay unveiled that betacyanins interact with BSA endothermically and spontaneously, with hydrophobic forces playing a dominant role. Remarkably, phyllocactin demonstrated higher binding affinity and stability to BSA than betanin. Furthermore, the incorporation of betacyanins into bread dose-dependently suppressed AGEs formation during baking and shows promise for inhibiting in vivo glycation process post-consumption. Overall, this study highlights the substantial potential of betacyanins as natural antiglycation agents.

RNA-Seq transcriptome profiling of immature grain wheat is a technique for understanding comparative modeling of baking quality.

Improving the baking quality is a primary challenge in the wheat flour production value chain, as baking quality represents a crucial factor in determining its overall value. In the present study, we conducted a comparative RNA-Seq analysis on the high baking quality mutant "O-64.1.10" genotype and its low baking quality wild type "Omid" cultivar to recognize potential genes associated with bread quality. The cDNA libraries were constructed from immature grains that were 15 days post-anthesis, with an average of 16.24 and 18.97 million paired-end short-read sequences in the mutant and wild-type, respectively. A total number of 733 transcripts with differential expression were identified, 585 genes up-regulated and 188 genes down-regulated in the "O-64.1.10" genotype compared to the "Omid". In addition, the families of HSF, bZIP, C2C2-Dof, B3-ARF, BES1, C3H, GRF, HB-HD-ZIP, PLATZ, MADS-MIKC, GARP-G2-like, NAC, OFP and TUB were appeared as the key transcription factors with specific expression in the "O-64.1.10" genotype. At the same time, pathways related to baking quality were identified through Kyoto Encyclopedia of Genes and Genomes. Collectively, we found that the endoplasmic network, metabolic pathways, secondary metabolite biosynthesis, hormone signaling pathway, B group vitamins, protein pathways, pathways associated with carbohydrate and fat metabolism, as well as the biosynthesis and metabolism of various amino acids, have a great deal of potential to play a significant role in the baking quality. Ultimately, the RNA-seq results were confirmed using quantitative Reverse Transcription PCR for some hub genes such as alpha-gliadin, low molecular weight glutenin subunit and terpene synthase (gibberellin) and as a resource for future study, 127 EST-SSR primers were generated using RNA-seq data.

Effect of magnetic fields on the structure, properties, baking performance of frozen wheat dough at different frozen stage.

As an emerging physical technology, magnetic fields have been used to improve the quality of frozen and refrigerated foods. This study compared the effect of applying a static magnetic field (2 mT) at different stages of freezing and storage on the quality of frozen dough. Results suggested that the magnetic field significantly impacted frozen dough quality. It not only prevented the formation of ice crystals during the pre-freezing stage but also inhibited ice crystal growth during the following frozen storage. This effect helped to maintain the integrity of gluten proteins and their adhesion to starch granules by preventing the breakage of disulfide bonds and the depolymerization of gluten macromolecules. It was also observed that yeast inactivation and glutathione release were reduced, resulting in improved air retention and air production capacity of the dough. This, in turn, led to a more appealing volume and texture quality of the finished bread.

Aerobic spore-forming bacteria associated with ropy bread: Identification, characterization and spoilage potential assessment.

Aerobic spore-forming (ASF) bacteria have been reported to cause ropiness in bread. Sticky and stringy degradation, discoloration, and an odor reminiscent of rotting fruit are typical characteristics of ropy bread spoilage. In addition to economic losses, ropy bread spoilage may lead to health risks, as virulent strains of ASF bacteria are not uncommon. However, the lack of systematic approaches to quantify physicochemical spoilage characteristics makes it extremely difficult to assess rope formation in bread. To address this problem, the aim of this study was to identify, characterize and objectively assess the spoilage potential of ASF bacteria associated with ropy bread. Hence, a set of 82 ASF bacteria, including isolates from raw materials and bakery environments as well as strains from international culture collections, were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and their species identity confirmed by 16S rRNA and gyrA or panC gene sequencing. A standardized approach supported by objective colorimetric measurements was developed to assess the rope-inducing potential (RIP) of a strain by inoculating autoclaved bread slices with bacterial spores. In addition, the presence of potential virulence factors such as swarming motility or hemolysis was investigated. This study adds B. velezensis, B. inaquosorum and B. spizizenii to the species potentially implicated of causing ropy bread spoilage. Most importantly, this study introduces a standardized classification protocol for assessing the RIP of a bacterial strain. Colorimetric measurements are used to objectively quantify the degree of breadcrumb discoloration. Furthermore, our results indicate that strains capable of inducing rope spoilage in bread often exhibit swarming motility and virulence factors such as hemolysis, raising important food quality considerations.

A critical evaluation on the valorization strategies to reduce and reuse orange waste in bakery industry.

Tons of orange by-products (OBPs) are generated during industrial orange processing. Currently, OBPs management is challenging due to their high amounts, physico-chemical characteristics (high water content, low pH, presence of essential oils) and seasonal nature of the production. Whereas agro-industrial OBPs can be highly valuable due to their abundant sources of bioactive compounds, which can add value to novel bakery products (e.g. bread, biscuits, cakes). This review covers the most recent research issues linked to the use of OBPs in bakery products, with a focus on available stabilization methods and on the main challenges to designing improved products. The application of OBPs improved the nutritional quality of bakery products, offering interesting sustainability benefits but also critical challenges. The valorization of OBPs may open new routes for the development of new natural ingredients for the food industry and lower food processing waste.

Review of formation mechanisms and quality regulation of chewiness in staple foods: Rice, noodles, potatoes and bread.

Staple foods serve as vital nutrient sources for the human body, and chewiness is an essential aspect of food texture. Age, specific preferences, and diminished eating functions have broadened the chewiness requirements for staple foods. Therefore, comprehending the formation mechanism of chewiness in staple foods and exploring approaches to modulate it becomes imperative. This article reviewed the formation mechanisms and quality control methods for chewiness in several of the most common staple foods (rice, noodles, potatoes and bread). It initially summarized the chewiness formation mechanisms under three distinct thermal processing methods: water medium, oil medium, and air medium processing. Subsequently, proposed some effective approaches for regulating chewiness based on mechanistic changes. Optimizing raw material composition, controlling processing conditions, and adopting innovative processing techniques can be utilized. Nonetheless, the precise adjustment of staple foods' chewiness remains a challenge due to their diversity and technical study limitations. Hence, further in-depth exploration of chewiness across different staple foods is warranted.

Croatian Action on Salt and Health (CRASH): On the Road to Success-Less Salt, More Health.

The World Health Organization recommends adjusting salt intake as a part of the nine global targets to reduce premature mortality from non-communicable chronic diseases as a priority and the most cost-effective intervention. In 2006, the main aim of the Croatian Action on Salt and Health was to decrease salt intake by 16% because of its critical intake and consequences on human health. We have organized educative activities to increase awareness on salt harmfulness, define food categories of prime interest, collaborate with industries and determine salt intake (24 h urine sodium excretion). It was determined that the proportion of salt in ready-to-eat baked bread should not exceed 1.4%. In the period 2014-2022, salt in semi-white bread was reduced by 14%, 22% in bakery and 25% in the largest meat industry. Awareness of the harmfulness of salt on health increased from 65.3% in 2008 to 96.9% in 2023 and salt intake was reduced by 15.9-1.8 g/day (22.8% men, 11.7% women). In the last 18 years, a significant decrease in salt intake was achieved in Croatia, awareness of its harmfulness increased, collaboration with the food industry was established and regulatory documents were launched. However, salt intake is still very high, underlying the need for continuation of efforts and even stronger activities.

Nitrogen use efficiency in bread wheat: Genetic variation and prospects for improvement.

Nitrogen (N) is one of the primary macronutrients required for crop growth and yield. This nutrient is especially limiting wheat yields in the dry and low fertile agro-ecologies having low N in the root zone soil strata. Moreover, majority of farmers in India and South Asia are small to marginal with meagre capacity to invest in costly nitrogen fertilizers. Therefore, there is an immense need to identify lines that use nitrogen efficiently. A set of 50 diverse wheat genotypes consisting of indigenous germplasm lines (05), cultivars released for commercial cultivation (23) and selected elite lines from CIMMYT nurseries (22) were evaluated in an alpha-lattice design with two replications, a six-rowed plot of 2.5m length for 24 agro morphological, physiological and NUE related traits during two consecutive crop seasons in an N-depleted precision field under two different N levels of 50%-N50 (T1) and 100%-N100 (T2) of recommended N, i.e., 100 kg/ha. Analysis of variance revealed significant genetic variation among genotypes for all the traits studied. About 11.36% yield reduction was observed at reduced N levels. Significant correlations among NUE traits and yield component traits were observed which indicated pivotal role of N remobilization to the grain in enhancing yield levels. Among N-insensitive genotypes identified based on their yielding ability at low N levels, UASBW13356, UASBW13358, UASBW13354, UASBW13357 and KRL1-4 showed their inherent genotypic plasticity toward N application. The genotypes with more yield and high to moderate NUtE can be used as parents for the breeding of N efficient genotypes for marginal agro-ecologies. Low N tolerant genotypes identified from the current investigation may be further utilized in the identification of genomic regions responsible for NUE and its deployment in wheat breeding programs. The comprehensive data of 24 traits under different nitrogen levels for diverse genotypes from India and global sources (mainly CIMMYT) should be useful for supporting breeding for NUE and thus will be of great help for small and marginal farmers in India and South Asia.

An SNP based genotyping assay for genes associated with drought tolerance in bread wheat.

BACKGROUND: SnRK2 plays vital role in responding to adverse abiotic stimuli. The applicability of TaSnRK2.4 and TaSnRK2.9 was investigated to leverage the potential of these genes in indigenous wheat breeding programs. METHODS: Genetic diversity was assessed using pre-existing markers for TaSnRK2.4 and TaSnRK2.9. Furthermore, new markers were also developed to enhance their broader applicability. KASP markers were designed for TaSnRK2.4, while CAPS-based markers were tailored for TaSnRK2.9. RESULTS: Analysis revealed lack of polymorphism in TaSnRK2.4 among Pakistani wheat germplasm under study. To validate this finding, available gel-based markers for TaSnRK2.4 were employed, producing consistent results and offering limited potential for application in marker-assisted wheat breeding with Pakistani wheat material. For TaSnRK2.9-5A, CAPS2.9-5A-1 and CAPS2.9-5A-2 markers were designed to target SNP positions at 308 nt and 1700 nt revealing four distinct haplotypes. Association analysis highlighted the significance of Hap-5A-1 of TaSnRK2.9-5A, which exhibited association with an increased number of productive tillers (NPT), grains per spike (GPS), and reduced plant height (PH) under well-watered (WW) conditions. Moreover, it showed positive influence on NPT under WW conditions, GPS under water-limited (WL) conditions, and PH under both WW and WL conditions. High selection intensity observed for Hap-5A-1 underscores the valuable role it has played in Pakistani wheat breeding programs. Gene expression studies of TaSnRK2.9-5A revealed the involvement of this gene in response to PEG, NaCl, low temperature and ABA treatments. CONCLUSION: These findings propose that TaSnRK2.9 can be effectively employed for improving wheat through marker-assisted selection in wheat breeding efforts.

Genetic correlation and Mendelian randomization analyses support causal relationships between dietary habits and age at menarche.

Dietary habits are essential in the mean age at menarche (AAM). However, the causal relationship between these factors remains unclear. Therefore, this study aimed to elucidate the genetic relationship between dietary habits and AAM. Genetic summary statistics for dietary habits were obtained from the UK Biobank. GWAS summary data for AAM was obtained from the ReproGen Consortium. Linkage disequilibrium score regression was used to test genetic correlations between dietary habits and AAM. The Mendelian randomization (MR) analyses used the inverse-variance weighted method. Genetic correlations with AAM were identified for 29 candi-date dietary habits, such as milk type (skimmed, semi-skimmed, full cream; coefficient = 0.2704, Pldsc = 1.13 × 10-14). MR evaluations revealed that 19 dietary habits were associated with AAM, including bread type (white vs. any other; OR 1.71, 95% CI 1.28-2.29, Pmr = 3.20 × 10-4), tablespoons of cooked vegetables (OR 0.437, 95% CI 0.29-0.67; Pmr = 1.30 × 10-4), and cups of coffee per day (OR 0.72, 95% CI 0.57-0.92, Pmr = 8.31 × 10-3). These results were observed to be stable under the sensitivity analysis. Our study provides potential insights into the genetic mechanisms underlying AAM and evidence that dietary habits are associated with AAM.

Effects of Green and Gold Kiwifruit Varieties on Antioxidant Neuroprotective Potential in Pigs as a Model for Human Adults.

Kiwifruit (KF) has shown neuroprotective potential in cell-based and rodent models by augmenting the capacity of endogenous antioxidant systems. This study aimed to determine whether KF consumption modulates the antioxidant capacity of plasma and brain tissue in growing pigs. Eighteen male pigs were divided equally into three groups: (1) bread, (2) bread + Actinidia deliciosa cv. 'Hayward' (green-fleshed), and (3) bread + A. chinensis cv. 'Hort16A' (yellow-fleshed). Following consumption of the diets for eight days, plasma and brain tissue (brain stem, corpus striatum, hippocampus, and prefrontal cortex) were collected and measured for biomarkers of antioxidant capacity, enzyme activity, and protein expression assessments. Green KF significantly increased ferric-reducing antioxidant potential (FRAP) in plasma and all brain regions compared with the bread-only diet. Gold KF increased plasma ascorbate concentration and trended towards reducing acetylcholinesterase activity in the brain compared with the bread-only diet. Pearson correlation analysis revealed a significant positive correlation between FRAP in the brain stem, prefrontal cortex, and hippocampus with the total polyphenol concentration of dietary interventions. These findings provide exploratory evidence for the benefits of KF constituents in augmenting the brain's antioxidant capacity that may support neurological homeostasis during oxidative stress.

Structural and Rheological Characterization of Vegetable Crispbread Enriched with Legume Purée.

Crispbread is gaining popularity as a healthy snack or bread substitute. This is a lightweight dry type of flat food that stays fresh for a very long time due to its lack of water and usually contains different types of grain flour, including gluten-containing wheat or rye flour. The incorporation of legume purée into crispbread represents an innovative approach to enhancing the nutritional profile and taste of the product. The rheological properties of various legume purées (chickpea, white bean, black bean, and red bean) mixed with citrus pectin were examined, revealing significant differences in fluid behavior and viscosity. Crispbread formulations were analyzed for water content and activity, color, structure, FT-IR spectra, water vapor adsorption isotherms, and sensory evaluation. The results showed the possibility of obtaining crispbread based on the purée of legumes and citrus pectin. Crispbread enriched with red bean purée exhibited low water activity (0.156) and water content (3.16%), along with a continuous porous structure, and received the highest sensory evaluation score among the products. These findings can be treated as a basis for the development of other innovative recipes and combinations using legumes.

Casein hydrolysate in naturally-fermented buckwheat sourdough: Effects on fermented and physicochemical characteristics, texture, and bacterial microbial composition.

The effect of a casein hydrolysate (CH) on the fermentation and quality of a naturally-fermented buckwheat sourdough (NFBS) were investigated, through assessing the fermentation characteristics, carbohydrate and protein degradation, texture, and bacterial composition of NFBS. According to the assaying data, CH might both increase the amount of lactic acid bacteria by 2.62 % and shorten the fermentation period by at least 3 h, subsequently leading to enhanced degradation of carbohydrate and protein, accompanied by a softer texture. More importantly, CH increased the relative abundance of lactobacillus in NFBS, making it the dominant bacterial genus and inhibited the growth of spoilage bacteria. In addition, Spearman correlation analysis indicated that the pH value, lactic and acetic acid contents, carbohydrates, protease activity, and these textural indices like hardness, elasticity, and adhesion had a positive/negative correlation with the bacterial composition of NFBS (Spearman correlation coefficient: -0.93-0.95). CH was thus regarded to be helpful to NFBS processing and production mainly by shortening its fermentation time, improving its fermentation performance, causing a finer texture and microstructure, and changing bacterial composition.

Insights into the hierarchical structure and physicochemical properties of starch isolated from fermented dough.

Understanding the hierarchical structure and physicochemical properties of starch isolated from fermented dough with different times (0-120 min) is valuable for improving the quality of fermented dough-based products. The results indicate that fermentation disrupted the starch granule surface and decreased the average particle size from 19.72 µm to 18.45 µm. Short-term fermentation (< 60 min) disrupted the crystalline, lamellar, short-range ordered molecular and helical structures of starch, while long-term fermentation (60-120 min) elevated the ordered degree of these structures. For example, relative crystallinity and double helix contents increased from 23.7 % to 26.8 % and 34.4 % to 37.2 %, respectively. During short-term fermentation, the structural amorphization facilitated interactions between starch molecular chains and water molecules, which increased the peak viscosity from 275.4 to 320.6 mPa·s and the swelling power from 7.99 to 8.52 g/g. In contrast, starches extracted from long-term fermented dough displayed the opposite results. Interestingly, the hardness and springiness of starch gels gradually decreased as fermentation time increased. These findings extend our understanding of the starch structure-property relationship during varied fermentation stages, potentially benefiting the production of better-fermented foods.

Novel chitosan-based smart bio-nanocomposite films incorporating TiO2 nanoparticles for white bread preservation.

Chitosan (CS)-based bio-nanocomposite food packaging films were prepared via solvent-casting method by incorporating a unique combination of additives and fillers, including polyvinyl alcohol (PVA), glycerol, Tween 80, castor oil (CO), and nano titanium dioxide (TiO2) in various proportions to enhance film properties. For a comprehensive analysis of the synthesized films, Fourier transform infrared (FTIR) spectroscopy, X-ray diffraction (XRD), thermogravimetric analysis (TGA), tensile testing, field emission scanning electron microscopy (FESEM), energy dispersive X-ray spectroscopy (EDS), and UV-vis spectrophotometry were employed. Furthermore, the antimicrobial efficacy of the films against S. aureus, E. coli, and A. niger was examined to assess their potential to preserve food from foodborne pathogens. The results claimed that the inclusion of castor oil and TiO2 nanoparticles considerably improved antimicrobial properties, UV-vis light barrier properties, thermal stability, optical transparency, and mechanical strength of the films, while reducing their water solubility, moisture content, water vapor and oxygen permeability. Based on the overall analysis, CS/PVA/CO/TiO2-0.3 film can be selected as the optimal one for practical applications. Furthermore, the practical application of the optimum film was evaluated using white bread as a model food product. The modified film successfully extended the shelf life of bread to 10 days, surpassing the performance of commercial LDPE packaging (6 days), and showed promising attributes for applications in the food packaging sector. These films exhibit superior antimicrobial properties, improved mechanical strength, and extended shelf life for food products, marking a sustainable and efficient alternative to conventional plastic packaging in both scientific research and industrial applications.

Improving the Quality of Wheat Flour Bread by a Thermophilic Xylanase with Ultra Activity and Stability Reconstructed by Ancestral Sequence and Computational-Aided Analysis.

Xylanase is an essential component used to hydrolyze the xylan in wheat flour to enhance the quality of bread. Presently, cold-activated xylanase is popularly utilized to aid in the development of dough. In this study, ancestral sequence reconstruction and molecular docking of xylanase and wheat xylan were used to enhance the activity and stability of a thermophilic xylanase. The results indicated that the ancestral enzyme TmxN3 exhibited significantly improved activity and thermal stability. The Vmax increased by 2.7 times, and the catalytic efficiency (Kcat/Km) increased by 1.7 times in comparison to TmxB. After being incubated at 100 °C for 120 min, it still retained 87.3% of its activity, and the half-life in 100 °C was 330 min, while the wild type xylanase was only 55 min. This resulted in an improved shelf life of bread, while adding TmxN3 considerably enhanced its quality with excellent volume and reduced hardness, chewiness, and gumminess. The results showed that the hardness was reduced by 55.2%, the chewiness was reduced by 40.11%, and the gumminess was reduced by 53.52%. To facilitate its industrial application, we further optimized the production conditions in a 5L bioreactor, and the xylanase activity reached 1.52 × 106 U/mL culture.