The role of branched-chain amino acids and their downstream metabolites in mediating insulin resistance.

Elevated levels of circulating branched-chain amino acids (BCAAs) and their associated metabolites have been strongly linked to insulin resistance and type 2 diabetes. Despite extensive research, the precise mechanisms linking increased BCAA levels with these conditions remain elusive. In this review, we highlight the key organs involved in maintaining BCAA homeostasis and discuss how obesity and insulin resistance disrupt the intricate interplay among these organs, thus affecting BCAA balance. Additionally, we outline recent research shedding light on the impact of tissue-specific or systemic modulation of BCAA metabolism on circulating BCAA levels, their metabolites, and insulin sensitivity, while also identifying specific knowledge gaps and areas requiring further investigation. Finally, we summarize the effects of BCAA supplementation or restriction on obesity and insulin sensitivity.

The roles of branched-chain amino acids metabolism in tumorigenesis and progression.

Branched-chain amino acids (BCAAs), including leucine, valine, and isoleucine, play crucial roles in regulating metabolic balance and maintaining physiological functions in the body. Extensive studies have been focused on their implications in obesity, diabetes, and cardiovascular diseases. Nevertheless, accumulating evidence suggests that BCAAs metabolism also plays significant roles in tumorigenesis and progression. In this review, we overview recent progress of the study on BCAAs metabolism including its relationship with epigenetic regulation. Particularly, we discuss the metabolic reprogramming and metabolic sensing of BCAAs and its intermediate metabolites in tumor cells and microenvironment to decipher their functions. An enhanced understanding of the roles and mechanism of BCAAs metabolism in tumorigenesis and progression will contribute to development of novel therapeutic strategies against tumor.

Molecular and physiological characterization of AIP1, encoding the acetolactate synthase regulatory subunit in rice.

Flooding deprives plants of oxygen and thereby causes severe stress by interfering with energy production, leading to growth retardation. Enzymes and metabolites may help protect plants from waterlogging and hypoxic environmental conditions. Acetolactate synthase (ALS) is a key enzyme in the biosynthesis of branched-chain amino acids (BCAAs), providing the building blocks for proteins and various secondary metabolites. Additionally, under energy-poor conditions, free BCAAs can be used as an alternative energy source by mitochondria through a catabolic enzyme chain reaction. In this study, we characterized ALS-INTERACTING PROTEIN 1 (OsAIP1), which encodes the regulatory subunit of ALS in rice (Oryza sativa). This gene was expressed in all parts of the rice plant, and its expression level was significantly higher in submerged and low-oxygen environments. Rice transformants overexpressing OsAIP1 showed a higher survival rate under hypoxic stress than did non-transgenic control plants under the same conditions. The OsAIP1-overexpressing plants accumulated increased levels of BCAAs, demonstrating that OsAIP1 is an important factor in the hypoxia resistance mechanism. These results suggest that ALS proteins are part of a defense mechanism that improves the tolerance of plants to low-oxygen environments.

Effects of Wheat Biscuits Enriched with Plant Proteins Incorporated into an Energy-Restricted Dietary Plan on Postprandial Metabolic Responses of Women with Overweight/Obesity.

This study investigates the effect of daily consumption of wheat biscuits enriched with plant proteins in postprandial metabolic responses of women with overweight/obesity who follow an energy-restricted diet. Thirty apparently healthy women participated in a 12-week randomized controlled trial and were assigned either to a control (CB) or an intervention (PB) group. Participants consumed daily either a conventional (CB) or an isocaloric wheat biscuit enriched with plant proteins (PB) containing high amounts of amino acids with appetite-regulating properties, i.e., BCAAs and L-arg. At baseline and the end of the intervention, a mixed meal tolerance test was performed. The responses of glucose, insulin, ghrelin, GLP-1, and glicentin were evaluated over 180 min. After 12 weeks, both groups experienced significant decreases in body weight, fat mass, and waist circumference. In the PB group, a trend towards higher weight loss was observed, accompanied by lower carbohydrate, fat, and energy intakes (p < 0.05 compared to baseline and CB group), while decreases in fasting insulin and the HOMA-IR index were also observed (p < 0.05 compared to baseline). In both groups, similar postprandial glucose, ghrelin, and GLP-1 responses were detected, while iAUC for insulin was lower (p < 0.05). Interestingly, the iAUC of glicentin was greater in the PB group (p < 0.05 compared to baseline). Subjective appetite ratings were beneficially affected in both groups (p < 0.05). Consumption of wheat biscuits enriched in plant proteins contributed to greater weight loss, lower energy intake, and insulin resistance and had a positive impact on postprandial glicentin response, a peptide that can potentially predict long-term weight loss and decreased food intake.

Bergenin Alleviates Ulcerative Colitis By Decreasing Gut Commensal Bacteroides vulgatus-Mediated Elevated Branched-Chain Amino Acids.

Ulcerative colitis is closely associated with the dysregulation of gut microbiota. There is growing evidence that natural products may improve ulcerative colitis by regulating the gut microbiota. In this research, we demonstrated that bergenin, a naturally occurring isocoumarin, significantly ameliorates colitis symptoms in dextran sulfate sodium (DSS)-induced mice. Transcriptomic analysis and Caco-2 cell assays revealed that bergenin could ameliorate ulcerative colitis by inhibiting TLR4 and regulating NF-κB and mTOR phosphorylation. 16S rRNA sequencing and metabolomics analyses revealed that bergenin could improve gut microbiota dysbiosis by decreasing branched-chain amino acid (BCAA) levels. BCAA intervention mediated the mTOR/p70S6K signaling pathway to exacerbate the symptoms of ulcerative colitis in mice. Notably, bergenin greatly decreased the symbiotic bacteria Bacteroides vulgatus (B. vulgatus), and the gavage of B. vulgatus increased BCAA concentrations and aggravated the symptoms of ulcerative colitis in mice. Our findings suggest that gut microbiota-mediated BCAA metabolism plays a vital role in the protective effect of bergenin on ulcerative colitis, providing novel insights for ulcerative colitis prevention through manipulation of the gut microbiota.

The Role of Branched-chain Amino Acids and Their Metabolism in Cardiovascular Diseases.

Branched-chain amino acids (BCAAs), including leucine, isoleucine, and valine, are essential amino acids for protein synthesis. Recent studies have yielded new insights into their diverse physiological and pathological roles in health and disease. Cardiovascular diseases (CVDs) are the leading cause of morbidity and mortality globally. An increasing number of clinical studies have demonstrated that high levels of circulating BCAAs are associated with an increased risk of CVDs. Animal studies have provided preliminary evidence linking BCAA intake and metabolism with cardiovascular diseases. Despite these insights, the causal relationship between BCAA metabolism and CVD remains poorly established, and the underlying mechanisms remain incompletely understood. Here, we aim to provide an update on the current understanding of the roles of BCAAs and their metabolism in the development and progression of various CVDs. We also discuss the potential strategies targeting BCAA nutrition and metabolism for the prevention and treatment of CVDs.

Effects of calorie, protein, and branched chain amino acid restriction on ovarian aging in mice.

Calorie restriction (CR) is an intervention that promotes longevity and preserves the ovarian reserve. Some studies have observed that the positive impacts of CR can be linked to restriction of protein (PR) and branched-chain amino acids (BCAAs) independent of calorie intake. The aim of this study was to compare the effects of protein and BCAA restriction to 30% CR on the ovarian reserve of female mice. For this, 3 month-old C57BL/6 female mice (n = 35) were randomized into four groups for four months dietary interventions including: control group (CTL; n = 8), 30% CR (CR; n = 9), protein restriction (PR; n = 9) and BCAA restriction (BCAAR; n = 9). Body mass gain, body composition, food intake, serum levels of BCAAs, ovarian reserve and estrous cyclicity were evaluated. We observed that CR, protein and BCAA restriction prevented weight gain and changed body composition compared to the CTL group. The BCAA restriction did not affect the ovarian reserve, while both PR and CR prevented activation of primordial follicles. This prevention occurred in PR group despite the lack of reduction of calorie intake compared to CTL group, and CR did not reduce protein intake in levels similar to the PR group. BCAA restriction resulted in increased calorie intake compared to CTL and PR mice, but only PR reduced serum BCAA levels compared to the CTL group. Our data indicates that PR has similar effects to CR on the ovarian reserve, whereas BCAA restriction alone did not affect it.

Amino acid profiles: exploring their diagnostic and pathophysiological significance in hypertension.

Hypertension, a major contributor to cardiovascular morbidity, is closely linked to amino acid metabolism. Amino acids, particularly branched-chain amino acids (BCAAs) and aromatic amino acids (AAAs), may play pivotal roles in the pathogenesis and potential management of hypertension. This review investigated the relationships between amino acid profiles, specifically BCAAs and AAAs, and hypertension, and examined their potential as diagnostic and therapeutic targets. An in-depth analysis was conducted on studies highlighting the associations of specific amino acids such as arginine, glycine, proline, glutamine, and the BCAAs and AAAs with hypertension. BCAAs and AAAs, alongside other amino acids like arginine, glycine, and proline, showed significant correlations with hypertension. These amino acids influence multiple pathways including nitric oxide synthesis, vascular remodeling, and neurotransmitter production, among others. Distinct amino acid profiles were discerned between hypertensive and non-hypertensive individuals. Amino acid profiling, particularly the levels of BCAAs and AAAs, offers promising avenues in the diagnostic and therapeutic strategies for hypertension. Future studies are crucial to confirm these findings and to delineate amino acid-based interventions for hypertension treatment.

Sex Differences in Branched-chain Amino Acid and Tryptophan Metabolism and Pathogenesis of Youth-onset Type 2 Diabetes.

OBJECTIVES: Insulin resistance is associated with elevations in plasma branched-chain amino acids (BCAAs). BCAAs compete with aromatic amino acids including tryptophan for uptake into ß cells. To explore relationships between BCAAs and tryptophan metabolism, adiposity, and glucose tolerance, we compared urine metabolites in overweight/obese youth with type 2 diabetes (T2D) with those in nondiabetic overweight/obese and lean youth. METHODS: Metabolites were measured in 24-hour and first-morning urine samples of 56 nondiabetic adolescents with overweight/obesity, 42 adolescents with T2D, and 43 lean controls, aged 12 to 21 years. Group differences were assessed by Kruskal Wallis or ANOVA. RESULTS: Groups were comparable for age, pubertal status, and ethnicity. Youth with T2D were predominantly female and had highest percent body fat. BCAAs, branched-chain ketoacids (BCKAs), tryptophan, and kynurenine were higher in urine of subjects with T2D. There were no differences between lean controls and nondiabetic youth with overweight/obesity. T2D was associated with diversion of tryptophan from the serotonin to the kynurenine pathway, with higher urinary kynurenine/serotonin ratio and lower serotonin/tryptophan and 5-HIAA/kynurenine ratios. Urinary BCAAs, BCKAs, tryptophan, and ratios reflecting diversion to the kynurenine pathway correlated positively with metrics of body fat and hemoglobin A1c. Increases in these metabolites in the obese T2D group were more pronounced and statistically significant only in adolescent girls. CONCLUSION: Increases in urinary BCAAs and BCKAs in adolescent females with T2D are accompanied by diversion of tryptophan metabolism from the serotonin to the kynurenine pathway. These adaptations associate with higher risks of T2D in obese adolescent females than adolescent males.

Branched-Chain Amino Acid Assembly into Amyloid-like Fibrils Provides a New Paradigm for Maple Syrup Urine Disease Pathology.

Inborn error of metabolism disorders (IEMs) are a family of diseases resulting from single-gene mutations that lead to the accumulation of metabolites that are usually toxic or interfere with normal cell function. The etiological link between metabolic alteration and the symptoms of IEMs is still elusive. Several metabolites, which accumulate in IEMs, were shown to self-assemble to form ordered structures. These structures display the same biophysical, biochemical, and biological characteristics as proteinaceous amyloid fibrils. Here, we have demonstrated, for the first time, the ability of each of the branched-chain amino acids (BCAAs) that accumulate in maple syrup urine disease (MSUD) to self-assemble into amyloid-like fibrils depicted by characteristic morphology, binding to indicative amyloid-specific dyes and dose-dependent cytotoxicity by a late apoptosis mechanism. We could also detect the presence of the assemblies in living cells. In addition, by employing several in vitro techniques, we demonstrated the ability of known polyphenols to inhibit the formation of the BCAA fibrils. Our study implies that BCAAs possess a pathological role in MSUD, extends the paradigm-shifting concept regarding the toxicity of metabolite amyloid-like structures, and suggests new pathological targets that may lead to highly needed novel therapeutic opportunities for this orphan disease.

Association Between Serum 3-Hydroxyisobutyric Acid and Prognosis in Patients With Chronic Heart Failure　- An Analysis of the KUNIUMI Registry Chronic Cohort.

BACKGROUND: Diabetes increases the risk of heart failure (HF). 3-Hydroxyisobutyric acid (3-HIB) is a muscle-derived metabolite reflecting systemic insulin resistance. In this study, we investigated the prognostic impact of 3-HIB in patients with chronic HF.Methods and Results: The KUNIUMI Registry chronic cohort is a community-based cohort study of chronic HF in Awaji Island, Japan. We analyzed the association between serum 3-HIB concentrations and adverse cardiovascular (CV) events in 784 patients from this cohort. Serum 3-HIB concentrations were significantly higher in patients with than without diabetes (P=0.0229) and were positively correlated with several metabolic parameters. According to Kaplan-Meier analysis, rates of CV death and HF hospitalization at 2 years were significantly higher among HF patients without diabetes in the high 3-HIB group (3-HIB concentrations above the median; i.e., >11.30 µmol/L) than in the low 3-HIB group (log-rank P=0.0151 and P=0.0344, respectively). Multivariable Cox proportional hazard models adjusted for established risk factors for HF revealed high 3-HIB as an independent predictor of CV death (hazard ratio [HR] 1.82; 95% confidence interval [CI] 1.16-2.85; P=0.009) and HF hospitalization (HR 1.72; 95% CI 1.17-2.53, P=0.006) in HF patients without diabetes, whereas no such trend was seen in subjects with diabetes. CONCLUSIONS: In a community cohort, circulating 3-HIB concentrations were associated with prognosis in chronic HF patients without diabetes.

Use of Branched-Chain Amino Acids as a Potential Treatment for Improving Nutrition-Related Outcomes in Advanced Chronic Liver Disease.

Advanced chronic liver disease (ACLD) represents a complex and multifactorial clinical entity characterized by liver dysfunction and associated complications. In recent years, the significance of nutritional status in ACLD prognosis has gained considerable attention. This review article delves into the multifactorial pathogenesis of malnutrition in ACLD and its profound consequences for health outcomes. We explore the clinical implications of secondary sarcopenia in ACLD and highlight the critical relevance of frailty in both decompensated and compensated ACLD. A specific focus of this review revolves around branched-chain amino acids (BCAAs) and their pivotal role in managing liver disease. We dissect the intricate relationship between low Fischer's ratio and BCAA metabolism in ACLD, shedding light on the molecular mechanisms involved. Furthermore, we critically evaluate the existing evidence regarding the effects of BCAA supplementation on outcomes in ACLD patients, examining their potential to ameliorate the nutritional deficiencies and associated complications in this population.

Resistant starch decreases intrahepatic triglycerides in patients with NAFLD via gut microbiome alterations.

Non-alcoholic fatty liver disease (NAFLD) is a hepatic manifestation of metabolic dysfunction for which effective interventions are lacking. To investigate the effects of resistant starch (RS) as a microbiota-directed dietary supplement for NAFLD treatment, we coupled a 4-month randomized placebo-controlled clinical trial in individuals with NAFLD (ChiCTR-IOR-15007519) with metagenomics and metabolomics analysis. Relative to the control (n = 97), the RS intervention (n = 99) resulted in a 9.08% absolute reduction of intrahepatic triglyceride content (IHTC), which was 5.89% after adjusting for weight loss. Serum branched-chain amino acids (BCAAs) and gut microbial species, in particular Bacteroides stercoris, significantly correlated with IHTC and liver enzymes and were reduced by RS. Multi-omics integrative analyses revealed the interplay among gut microbiota changes, BCAA availability, and hepatic steatosis, with causality supported by fecal microbiota transplantation and monocolonization in mice. Thus, RS dietary supplementation might be a strategy for managing NAFLD by altering gut microbiota composition and functionality.

Depiction of Branched-Chain Amino Acids (BCAAs) in Diabetes with a Focus on Diabetic Microvascular Complications.

Type 2 diabetes mellitus (T2DM) still holds the title as one of the most debilitating chronic diseases with rising prevalence and incidence, including its complications such as retinal, renal, and peripheral nerve disease. In order to develop novel molecules for diagnosis and treatment, a deep understanding of the complex molecular pathways is imperative. Currently, the existing agents for T2DM treatment target only blood glucose levels. Over the past decades, specific building blocks of proteins-branched-chain amino acids (BCAAs) including leucine, isoleucine, and valine-have gained attention because they are linked with insulin resistance, pre-diabetes, and diabetes development. In this review, we discuss the hypothetical link between BCAA metabolism, insulin resistance, T2DM, and its microvascular complications including diabetic retinopathy and diabetic nephropathy. Further research on these amino acids and their derivates may eventually pave the way to novel biomarkers or therapeutic concepts for the treatment of diabetes and its accompanied complications.

Metabolic Profiling in Tuberous Roots of Ranunculus asiaticus L. as Influenced by Vernalization Procedure.

Ranunculus asiaticus L. is an ornamental geophyte. In commercial practice, it is mainly propagated by rehydrated tuberous roots. Vernalization before planting is a common practice to overcome the natural dormancy of tuberous roots; however, little is known about the mechanisms underlying the plant's response to low temperatures. We investigated the influence of three preparation procedures of tuberous roots, only rehydration (control, C), and rehydration plus vernalization at 3.5 °C for 2 weeks (V2) and for 4 weeks (V4), on plant growth, leaf photosynthesis, flowering, and metabolism in plants of two hybrids, MBO (early flowering, pale orange flower) and MDR (medium earliness, bright orange flower), grown in pots in an unheated greenhouse. We reported the responses observed in the aerial part in a previous article in this journal. In this paper, we show changes in the underground organs in carbohydrate, amino acids, polyphenols, and protein levels throughout the growing cycle in the different plant stages: pre-planting, vegetative growth, and flowering. The metabolic profile revealed that the two hybrids had different responses to the root preparation procedure. In particular, MBO synthesized GABA and alanine after 2 weeks and sucrose after 4 weeks of vernalization. In contrast, MDR was more sensitive to vernalization; in fact, a higher synthesis of polyphenols was observed. However, both hybrids synthesized metabolites that could withstand exposure to low temperatures.

Expression Pattern and Prognostic Analysis of Branched-Chain Amino Acid Catabolism-Related Genes in Non-Small Cell Lung Cancer.

BACKGROUND: The purpose of our study is to analyze the expression pattern and prognostic value of catabolism-related enzymes of branched-chain amino acids (BCAAs) in non-small cell lung cancer (NSCLC). METHODS: Differential expression analysis, mutation, copy number variation (CNV), methylation analysis, and survival analysis of BCAAs catabolism-related enzymes in NSCLC were performed using the Cancer Genome Atlas (TCGA) database. RESULTS: Six and seven differentially expressed genes were obtained in lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LUSC), respectively. IL4I1 was located at the core regulatory nodes in the gene co-expression networks of both LUAD and LUSC. The AOX1 mutation rate was the highest in both LUAD and LUSC. For CNV, IL4I1 was up-regulated in both LUAD and LUSC with an increase in copy number, whereas AOX1 and ALDH2 were differentially regulated in the two subtypes of lung cancer. In patients with NSCLC, high expression of IL4I1 was associated with lower overall survival (OS), and low expression of ALDH2 predicted shorter disease-free survival (DFS). ALDH2 expression was related with LUSC survival. CONCLUSIONS: This study explored the biomarkers of BCAAs catabolism related to the prognosis of NSCLC, which provided a theoretical foundation to guide the clinical diagnosis and treatment of NSCLC.

Identification of candidate metabolite biomarkers for metabolic syndrome and its five components in population-based human cohorts.

BACKGROUND: Metabolic Syndrome (MetS) is characterized by risk factors such as abdominal obesity, hypertriglyceridemia, low high-density lipoprotein cholesterol (HDL-C), hypertension, and hyperglycemia, which contribute to the development of cardiovascular disease and type 2 diabetes. Here, we aim to identify candidate metabolite biomarkers of MetS and its associated risk factors to better understand the complex interplay of underlying signaling pathways. METHODS: We quantified serum samples of the KORA F4 study participants (N = 2815) and analyzed 121 metabolites. Multiple regression models adjusted for clinical and lifestyle covariates were used to identify metabolites that were Bonferroni significantly associated with MetS. These findings were replicated in the SHIP-TREND-0 study (N = 988) and further analyzed for the association of replicated metabolites with the five components of MetS. Database-driven networks of the identified metabolites and their interacting enzymes were also constructed. RESULTS: We identified and replicated 56 MetS-specific metabolites: 13 were positively associated (e.g., Val, Leu/Ile, Phe, and Tyr), and 43 were negatively associated (e.g., Gly, Ser, and 40 lipids). Moreover, the majority (89%) and minority (23%) of MetS-specific metabolites were associated with low HDL-C and hypertension, respectively. One lipid, lysoPC a C18:2, was negatively associated with MetS and all of its five components, indicating that individuals with MetS and each of the risk factors had lower concentrations of lysoPC a C18:2 compared to corresponding controls. Our metabolic networks elucidated these observations by revealing impaired catabolism of branched-chain and aromatic amino acids, as well as accelerated Gly catabolism. CONCLUSION: Our identified candidate metabolite biomarkers are associated with the pathophysiology of MetS and its risk factors. They could facilitate the development of therapeutic strategies to prevent type 2 diabetes and cardiovascular disease. For instance, elevated levels of lysoPC a C18:2 may protect MetS and its five risk components. More in-depth studies are necessary to determine the mechanism of key metabolites in the MetS pathophysiology.

Preliminary Study on the Association of Serum Branched-Chain Amino Acids With Lipid and Hepatic Markers.

Background: Serum levels of branched-chain amino acids (BCAAs) are associated with various vital physiological functions and thus elevation in circulating levels results in several metabolic disturbances. Serum levels of BCAAs are strong predictors of various metabolic disorders. Their association with cardiovascular health is uncertain. The study aimed to investigate the association of BCAAs with circulating levels of vital cardiovascular and hepatic markers. Methods: The study population of 714 individuals was included from the population tested for the vital cardio and hepatic biomarkers at the Vibrant America Clinical Laboratories. The subjects were stratified into four quartiles based on the serum levels of BCAAs, and their association with vital markers was studied using the Kruskal-Wallis test. Pearson's correlation analyzed the univariant relationship of BCAAs with selected cardio and hepatic markers. Results: BCAAs exhibited a strong negative correlation with serum HDL. Serum triglycerides were found to have a positive correlation with serum levels of leucine and valine. Univariant analysis exhibited a strong negative correlation between serum levels of BCAAs and HDL, and a positive correlation was observed between triglycerides and amino acids isoleucine and leucine. Among analyzed hepatic markers, alanine transaminase exhibited a considerable association with BCAAs. Conclusions: The elevated levels of serum BCAAs are strongly associated with serum HDL and triglycerides. Consumption of these supplements must be in coordination with healthcare providers to avoid metabolic and cardiovascular risk.

Identification and Characterization of an Antifungal Gene Mt1 from Bacillus subtilis by Affecting Amino Acid Metabolism in Fusarium graminearum.

Fusarium head blight is a devastating disease that causes significant economic losses worldwide. Fusarium graminearum is a crucial pathogen that requires close attention when controlling wheat diseases. Here, we aimed to identify genes and proteins that could confer resistance to F. graminearum. By extensively screening recombinants, we identified an antifungal gene, Mt1 (240 bp), from Bacillus subtilis 330-2. We recombinantly expressed Mt1 in F. graminearum and observed a substantial reduction in the production of aerial mycelium, mycelial growth rate, biomass, and pathogenicity. However, recombinant mycelium and spore morphology remained unchanged. Transcriptome analysis of the recombinants revealed significant down-regulation of genes related to amino acid metabolism and degradation pathways. This finding indicated that Mt1 inhibited amino acid metabolism, leading to limited mycelial growth and, thus, reduced pathogenicity. Based on the results of recombinant phenotypes and transcriptome analysis, we hypothesize that the effect of Mt1 on F. graminearum could be related to the metabolism of branched-chain amino acids (BCAAs), the most affected metabolic pathway with significant down-regulation of several genes. Our findings provide new insights into antifungal gene research and offer promising targets for developing novel strategies to control Fusarium head blight in wheat.

Modified whey protein isolate gel prepared by thermal aggregation combined with transglutaminase crosslinking achieves Casein-like slow digestion in vitro and in vivo.

Our study aimed to fabricate a modified slow-digestive whey protein isolate (WPI), which can supply enough branched-chain amino acids (BCAAs) during long-term fasting. The WPI aqueous solution (10 % w/v) was treated by heat (80 ℃) to unfold the protein tertiary structure, and subsequently treated with transglutaminase to form a gel via cross-linking. The powder of the WPI gel was obtained by spray drying, which can dissolve in water easily and self-assemble into gels again. This modified WPI contained protein aggregates with high molecular weight, and kept a stable gel-like structure under simulated gastric digestion conditions (pH = 3, 37 ℃). A dense honeycomb internal microstructure of the freeze-dried gel was observed. Further, we found that the WPI gel successfully achieved a casein-like digestible ratio (37.37 %) and released more BCAAs (0.18 mg/mL) than casein during the 4 h of in vitro simulated digestion based on the INFOGEST method. Finally, our results showed that the C57BL/6 mice oral administrated with the modified WPI gel had consistently higher BCAAs concentration (0.052 mg/mL) in their blood serum than the mice with normal WPI intake during the 6 h of in vivo digestion.