Papaya, which is mainly cultivated in the southeastern region of China, is one of the four famous fruits in Lingnan. It is favored by people because of its edible and medicinal value. Fructose-6-phosphate, 2-kinase/fructose-2, 6-bisphosphatase (F2KP) is a unique bifunctional enzyme with a kinase domain and an esterase domain that catalyzes the synthesis and degradation of fructose-2, 6-bisphosphate (Fru-2, 6-P₂), an important regulator of glucose metabolism in organisms. In order to study the function of the gene CpF2KP encoding the enzyme in papaya, it is particularly important to obtain the target protein. In this study, the coding sequence (CDS) of CpF2KP, with a full-length of 2 274 bp, was got from the papaya genome. The amplified sequence of full-length CDS was cloned into the vector PGEX-4T-1 which was double digested with EcoR I and BamH I. The amplified sequence was constructed into a prokaryotic expression vector by genetic recombination. After exploring the induction conditions, the results of SDS-PAGE showed that the size of the recombinant GST-CpF2KP protein was about 110 kDa. The optimum IPTG concentration and temperature for CpF2KP induction were 0.5 mmol/L and 28 ℃, respectively. The purified sin[A1] gle target protein was obtained after purifying the induced CpF2KP protein. In addition, the expression level of this gene was detected in different tissues, and showed that the gene was expressed at the highest level in seeds and the lowest in pulp. This study provides an important basis for further revealing the function of CpF2KP protein and studying the involved biological processes of this gene in papaya.
Humans ; Carica/genetics* ; Recombinant Proteins ; Carbohydrate Metabolism ; Cloning, Molecular ; China

As specialized intracellular parasite, viruses have no ability to metabolize independently, so they completely depend on the metabolic mechanism of host cells. Viruses use the energy and precursors provided by the metabolic network of the host cells to drive their replication, assembly and release. Namely, viruses hijack the host cells metabolism to achieve their own replication and proliferation. In addition, viruses can also affect host cell metabolism by the expression of auxiliary metabolic genes (AMGs), affecting carbon, nitrogen, phosphorus, and sulfur cycles, and participate in microbial-driven biogeochemical cycling. This review summarizes the effect of viral infection on the host's core metabolic pathway from four aspects: cellular glucose metabolism, glutamine metabolism, fatty acid metabolism, and viral AMGs on host metabolism. It may facilitate in-depth understanding of virus-host interactions, and provide a theoretical basis for the treatment of viral diseases through metabolic intervention.
Humans ; Metabolic Networks and Pathways ; Virus Diseases ; Carbohydrate Metabolism ; Host Microbial Interactions ; Lipid Metabolism

Child ; Humans ; Carbohydrate Metabolism, Inborn Errors ; Microcephaly ; Psychomotor Disorders ; Seizures

Carbohydrate Epimerases/metabolism* ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Esophageal Neoplasms/genetics* ; Gene Expression Regulation, Neoplastic ; Humans ; Ketone Oxidoreductases/metabolism* ; MAP Kinase Signaling System ; Matrix Metalloproteinase 2/genetics* ; Matrix Metalloproteinase 9 ; Neoplasm Invasiveness/genetics*

OBJECTIVE: To analyze the clinical phenotype and variant of SLC2A1 gene in a Chinese pedigree affected with glucose transporter type 1 deficiency syndrome (GLUT1-DS). METHODS: Clinical data of a child who was treated due to delayed motor and language development and his family members were collected. DNA was extracted from peripheral blood samples and subjected to high-throughput medical exome sequencing. Candidate variant was verified by Sanger sequencing of his parents and sister. The genotype-phenotype correlation was explored. RESULTS: The child, his mother and sister had common manifestations such as delayed mental and motor development, poor exercise tolerance, easy fatigue and paroxysmal dystonia, but the difference was that the child and his mother had microcephaly and seizures, while his sister did not. A heterozygous missense SLC2A1 c.191T>C (p.L64P) variant was identified in all affected members, which was unreported previously. CONCLUSION The missense SLC2A1 c.191T>C (p.L64P) variant probably underlay the disease in the proband and his mother and sister. Variability of the clinical phenotypes has reflected the genetic and phenotypic diversity of GLUT1-DS. Detection of the novel variant has enriched the spectrum of GLUT1-DS mutations.
Carbohydrate Metabolism, Inborn Errors ; China ; Glucose Transporter Type 1/genetics* ; Humans ; Monosaccharide Transport Proteins/deficiency* ; Mutation ; Pedigree ; Phenotype

Sucrose is a natural product occurs widely in nature. In living organisms such as plants, sucrose phosphate synthase (SPS) is the key rate-limiting enzyme for sucrose synthesis. SPS catalyzes the synthesis of sucrose-6-phosphate, which is further hydrolyzed by sucrose phosphatase to form sucrose. Researches on SPS in recent decades have been focused on the determination of enzymatic activity of SPS, the identification of the inhibitors and activators of SPS, the covalent modification of SPS, the carbohydrate distribution in plants regulated by SPS, the mechanism for promoting plant growth by SPS, the sweetness of fruit controlled by SPS, and many others. A systematic review of these aspects as well as the crystal structure and catalytic mechanism of SPS are presented.
Carbohydrate Metabolism ; Glucosyltransferases/metabolism* ; Plants/metabolism* ; Sucrose

Lifestyle interventions, including dietary adjustments and exercise, are important for obesity management. This study enrolled adults with overweight or obesity to explore whether either low-carbohydrate diet (LCD) or exercise is more effective in metabolism improvement. Forty-five eligible subjects were randomly divided into an LCD group (n = 22) and an exercise group (EX, n = 23). The subjects either adopted LCD (carbohydrate intake < 50 g/day) or performed moderate-to-vigorous exercise (⩾ 30 min/day) for 3 weeks. After the interventions, LCD led to a larger weight loss than EX ( - 3.56 ± 0.37 kg vs. - 1.24 ± 0.39 kg, P < 0.001), as well as a larger reduction in fat mass ( - 2.10 ± 0.18 kg vs. - 1.25 ± 0.24 kg, P = 0.007) and waist circumference ( - 5.25 ± 0.52 cm vs. - 3.45 ± 0.38 cm, P = 0.008). Both interventions reduced visceral and subcutaneous fat and improved liver steatosis and insulin resistance. Triglycerides decreased in both two groups, whereas low-density lipoprotein cholesterol increased in the LCD group but decreased in the EX group. Various glycemic parameters, including serum glycated albumin, mean sensor glucose, coefficient of variability (CV), and largest amplitude of glycemic excursions, substantially declined in the LCD group. Only CV slightly decreased after exercise. This pilot study suggested that the effects of LCD and exercise are similar in alleviating liver steatosis and insulin resistance. Compared with exercise, LCD might be more efficient for weight loss and glucose homeostasis in people with obesity.
Adult ; Blood Glucose ; Diet, Carbohydrate-Restricted ; Homeostasis ; Humans ; Pilot Projects ; Weight Loss

OBJECTIVE: To explore the genetic basis for a child with ocular anomaly, microcephaly, growth retardation and intrauterine growth restriction. METHODS: The patient underwent ophthalmologic examinations including anterior segment photography, fundus color photography, and fundus fluorescein angiography. The patient and her parents were subjected to whole exome sequencing. Candidate variants were verified by Sanger sequencing and bioinformatic analysis. RESULTS: The patient was found to have bilateral persistent pupillary membrane and coloboma of inferior iris, in addition with macular dysplasia and radial pigmentation near the hemal arch of the temporal retina. She was found to have carried compound heterozygous missense variants of the PHGDH gene, namely c.196G>A and c.1177G>A, which were respectively inherited from her father and mother. Bioinformatic analysis suggested both variants to be pathogenic. CONCLUSION The patient was diagnosed with phosphoglycerate dehydrogenase deficiency. Above finding has enriched the phenotypic spectrum of the disease with ocular manifestations.
Carbohydrate Metabolism, Inborn Errors/genetics* ; Child ; Coloboma ; Female ; Humans ; Microcephaly/genetics* ; Mutation ; Phenotype ; Phosphoglycerate Dehydrogenase/genetics* ; Psychomotor Disorders/genetics* ; Seizures/genetics* ; Whole Exome Sequencing

D-allulose-3-epimerase (DPEase) is the key enzyme for isomerization of D-fructose to D-allulose. In order to improve its thermal stability, short amphiphilic peptides (SAP) were fused to the N-terminal of DPEase. SDS-PAGE analysis showed that the heterologously expressed DPEase folded correctly in Bacillus subtilis, and the protein size was 33 kDa. After incubation at 40 °C for 48 h, the residual enzyme activity of SAP1-DSDPEase was 58%. To make the recombinant B. subtilis strain reusable, cells were immobilized with a composite carrier of sodium alginate (SA) and titanium dioxide (TiO2). The results showed that 2% SA, 2% CaCl2, 0.03% glutaraldehyde solution and a ratio of TiO2 to SA of 1:4 were optimal for immobilization. Under these conditions, up to 82% of the activity of immobilized cells could be retained. Compared with free cells, the optimal reaction temperature of immobilized cells remained unchanged at 80 °C but the thermal stability improved. After 10 consecutive cycles, the mechanical strength remained unchanged, while 58% of the enzyme activity could be retained, with a conversion rate of 28.8% achieved. This study demonstrated a simple approach for using SAPs to improve the thermal stability of recombinant enzymes. Moreover, addition of TiO2 into SA during immobilization was demonstrated to increase the mechanical strength and reduce cell leakage.
Bacillus subtilis/metabolism* ; Carbohydrate Epimerases/genetics* ; Enzyme Stability ; Enzymes, Immobilized/metabolism* ; Fructose ; Hydrogen-Ion Concentration ; Racemases and Epimerases ; Temperature

Heparin and heparan sulfate are a class of glycosaminoglycans for clinical anticoagulation. Heparosan N-sulfate-glucuronate 5-epimerase (C5, EC 5.1.3.17) is a critical modifying enzyme in the synthesis of heparin and heparan sulfate, and catalyzes the inversion of carboxyl group at position 5 on D-glucuronic acid (D-GlcA) of N-sulfoheparosan to form L-iduronic acid (L-IdoA). In this study, the heparin C5 epimerase gene Glce from zebrafish was expressed and molecularly modified in Escherichia coli. After comparing three expression vectors of pET-20b (+), pET-28a (+) and pCold Ⅲ, C5 activity reached the highest ((1 873.61±5.42) U/L) with the vector pCold Ⅲ. Then we fused the solution-promoting label SET2 at the N-terminal for increasing the soluble expression of C5. As a result, the soluble protein expression was increased by 50% compared with the control, and the enzyme activity reached (2 409±6.43) U/L. Based on this, site-directed mutations near the substrate binding pocket were performed through rational design, the optimal mutant (V153R) enzyme activity and specific enzyme activity were (5 804±5.63) U/L and (145.1±2.33) U/mg, respectively 2.41-fold and 2.28-fold of the original enzyme. Modification and expression optimization of heparin C5 epimerase has laid the foundation for heparin enzymatic catalytic biosynthesis.
Animals ; Carbohydrate Epimerases ; biosynthesis ; chemistry ; genetics ; Escherichia coli ; Gene Expression ; Heparin ; metabolism ; Heparitin Sulfate ; metabolism ; Iduronic Acid ; metabolism ; Zebrafish Proteins ; biosynthesis ; chemistry ; genetics