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*

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

Early-onset progressive encephalopathy is a lethal encephalopathy caused by NAXE gene mutations. This paper reports the clinical and genetic features of a patient with early-onset progressive encephalopathy. A 4-year-old boy admitted to the hospital had repeated walking instability and limb weakness for 2 years. The patient and his elder brother (already dead) had clinical onset at 2 years of age. Both of them showed symptoms such as strabismus, ataxia, reduced muscle tone, delayed development, and repeated respiratory failure after infection. The NAXE gene of the patient showed new compound heterozygous mutations, i.e., c.255 (exon 2) A>T from his mother and c.361 (exon 3) G>A from his father. The NAXE gene encodes an epimerase that is essential for the repair of cellular metabolites of NADHX and NADPHX. This disease is associated with a deficiency of the mitochondrial NAD(P)HX repair system. Patients usually have rapid disease progression. They are also quite likely to have respiratory failure immediately after infection.
Adult ; Age of Onset ; Base Sequence ; Brain Diseases ; enzymology ; genetics ; Child, Preschool ; Disease Progression ; Female ; Heterozygote ; Humans ; Male ; Mutation ; Racemases and Epimerases ; genetics

OBJECTIVETo investigate the clinical manifestations, pathological characteristics, and treatments of urothelial-type mucinous adenocarcinoma of the prostate (UMAP).

METHODSWe reported a case of UMAP, reviewed relevant literature, and analyzed the clinicopaothological features, diagnosis, treatment, and prognosis of the disease.

RESULTSThe patient was a 60-year-old male and underwent transurethral resection of the prostate for dysuria. Postoperative pathology indicated mucinous adenocarcinoma and sigmoidoscopy revealed no primary colon cancer. Immunohistochemical staining showed the negative expressions of PSA and P504s and positive expressions of CK7, CK34 β E12, CK20, and CDX2. Thus UMAP was confirmed and treated by intensity-modulated radiotherapy. Then the patient was followed up for 30 months, which showed desirable therapeutic result, with neither local progression nor distant metastasis.

CONCLUSIONUMAP has a bad prognosis and its diagnosis depends on pathological and immunohistocchemical examinations. It responds well to radical prostatectomy but is not sensitive to endocrine therapy. Radiotherapy can be considered for those who are not fit to receive radical prostatectomy.

Adenocarcinoma, Mucinous ; metabolism ; pathology ; therapy ; Humans ; Keratins ; metabolism ; Male ; Middle Aged ; Neoplasm Proteins ; metabolism ; Prognosis ; Prostatectomy ; Prostatic Neoplasms ; metabolism ; pathology ; therapy ; Racemases and Epimerases ; metabolism

Taxol is one of the most potent anti-cancer agents, which is extracted from the plants of Taxus species. Isopentenyl diphosphate isomerase (IPI) catalyzes the reversible transformation between IPP and DMAPP, both of which are the general 5-carbon precursors for taxol biosynthesis. In the present study, a new gene encoding IPI was cloned from Taxus media (namely TmIPI with the GenBank Accession Number KP970677) for the first time. The full-length cDNA of TmIPI was 1 232 bps encoding a polypeptide with 233 amino acids, in which the conserved domain Nudix was found. Bioinformatic analysis indicated that the sequence of TmIPI was highly similar to those of other plant IPI proteins, and the phylogenetic analysis showed that there were two clades of plant IPI proteins, including IPIs of angiosperm plants and IPIs of gymnosperm plants. TmIPI belonged to the clade of gymnosperm plant IPIs, and this was consistent with the fact that Taxus media is a plant species of gymnosperm. Southern blotting analysis demonstrated that there was a gene family of IPI in Taxus media. Finally, functional verification was applied to identify the function of TmIPI. The results showed that biosynthesis of β-carotenoid was enhanced by overexpressing TmIPI in the engineered E. coli strain, and this suggested that TmIPI might be a key gene involved in isoprenoid/terpenoid biosynthesis.
Amino Acid Sequence ; Carbon-Carbon Double Bond Isomerases ; genetics ; Cloning, Molecular ; DNA, Complementary ; genetics ; Escherichia coli ; Paclitaxel ; biosynthesis ; Phylogeny ; Plant Proteins ; genetics ; Taxus ; enzymology ; genetics

Prostate cancer is one of the most common malignant tumors in the male urinary system as well as the second leading cause of cancer death in men. Prostate specific antigen (PSA) screening is the main method for the early diagnosis of prostate cancer, but has a low specificity for its detection. In recent years, a variety of tumor markers with high sensitivity and specificity have been found. This review focuses on some of the more promising tumor biomarkers such as prostate cancer antigen 3, early prostate cancer antigen, prostate-specific membrane antigen, alpha-methylacyl-CoA racemase, and vascular endothelial growth factor.
Antigens, Neoplasm ; blood ; Antigens, Surface ; blood ; Biomarkers, Tumor ; blood ; Early Detection of Cancer ; GPI-Linked Proteins ; blood ; Glutamate Carboxypeptidase II ; blood ; Humans ; Male ; Neoplasm Proteins ; blood ; Prostate-Specific Antigen ; blood ; Prostatic Neoplasms ; diagnosis ; Racemases and Epimerases ; blood ; Sensitivity and Specificity ; Vascular Endothelial Growth Factor A ; blood

OBJECTIVETo study the clinicopathological features, differential diagnosis and prognosis of clear cell papillary renal cell carcinoma (CCPRCC).

METHODSThe histological, immunohistochemical, and molecular features were studied in 11 cases and follow-up data were also analyzed.

RESULTSThere were a total of 3 females and 8 males. The age of patients were ranged from 33 to 72 years(mean age 52.5 years). The diameters of tumors varied from 1cm to 4 cm. Histologically, papillary and cystic architecture were present at least focally in all tumors. The papillae were covered by small to medium-sized cuboidal cells with abundant clear cytoplasm and often showed extensive secondary branching, which were often folded and densely packed, resulting in a solid appearance. The nuclei were round and uniform in shape; nucleoli were not prominent (Fuhrman grade 1 or 2). Neither mitotic figures nor necrosis was present. All 11 cases exhibited moderate to strong positivity for CK7, CA9, vimentin, and HIF-1α, coupled with negative reactions for CD10, P504S, and TFE3. Ksp-cadherin was positively expressed in 8 cases.VHL gene mutations were not found in all 11 cases. Losses of chromosomes 3 (monoploid chromosome 3) was detected in 3 cases.

CONCLUSIONSCCPRCC is uncommon and seemed to be an indolent tumor. The differential diagnosis should be included tumors, which harbor clear cell and papillary structure including clear cell renal cell carcinoma, papillary renal cell carcinoma, Xp11 translocation renal cell carcinoma, and CCPRCC. Immunohistochemical and molecular analysis may be help for its diagnosis.

Adult ; Aged ; Carcinoma, Renal Cell ; chemistry ; genetics ; pathology ; ultrastructure ; Chromosomes, Human, Pair 3 ; Female ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; analysis ; Kidney Neoplasms ; chemistry ; genetics ; pathology ; ultrastructure ; Male ; Middle Aged ; Mutation ; Prognosis ; Racemases and Epimerases ; analysis ; Translocation, Genetic ; Tumor Burden

OBJECTIVETo investigate the clinicopathological characteristics and the diagnosis of multilocular cystic renal cell carcinoma (MCRCC).

METHODSThe clinicopathological data of 19 MCRCC cases were collected and immunohistochemical staining assays were carried out. Forty-six cases of other cystic kidney lesions within the same period were collected as controls, including extensively cystic clear cell RCC (12 cases), clear cell tubulopapillary renal cell carcinoma (6 cases), tubulocystic carcinoma (2 cases), simple cortical cysts (22 cases), multilocular cystic nephroma (1 cases) and multicystic kidney (3 cases).

RESULTSThe patients included 14 males and 5 females. The ages ranged from 31 to 66 years (median age = 50 years). Most of the MCRCC cases were detected incidentally in physical examination, occasionally accompanied with hematuria, back pain or other symptoms. The follow-up period of 17 patients ranged from 6 to 170 months. All patients were alive without evidence of tumor recurrence or metastasis. Pathological findings showed that macroscopically, tumor size ranges from 1.5 to 7.0 cm in the maximum diameter, generally a entirely of various sized. The cysts contain serous, hemorrhagic or turbid fluid. Solid areas or substantially discernible mural nodules were absent; histologicallly, single layer of cuboidal and flattened epithelial tumor cells were lined in the cysts, described as clear cytoplasm, small nuclear, no nucleoli and low Fuhrman nuclear grade (I or II). Multilayer tumor cells could be observed in a few cysts, with granular cytoplasm and small intracystic papillae formed. The clear tumor cell clusters, similar as cystic lined tumor cells, were seen within pathological fibrous in almost all cases, and significant myofibroblastic proliferation was found in 14 cases. Immunohistochemically, the cysts lined epithelial cells and the clear tumor cell clusters were positive for epithelium markers, including CKpan(19/19), EMA(16/19) and CK7 (15/19); higher percentage of CAIX (17/19) and PAX8(15/19) than control groups, but lower percentage of CD10 (7/19), RCC (6/19) and AMACR(2/19); and all were negative for 34βE12, CD117 and CD68.

CONCLUSIONSMultilocular cysts, clear cells clusters of low Fuhrman grade within fibrous septa and capillary vessel proliferation under epithelium are important features of MCRCC. The united using of CAIX, CK7, CD10 and RCC is helpful for differentiating variable cystic renal tumor. MCRCC usually has an excellent prognosis, nephron sparing surgery is first recommended as a therapeutic strategy.

Adenocarcinoma, Clear Cell ; metabolism ; pathology ; Biomarkers ; Carcinoma, Renal Cell ; metabolism ; pathology ; Cysts ; metabolism ; pathology ; Diagnosis, Differential ; Female ; Humans ; Kidney Diseases, Cystic ; metabolism ; pathology ; Kidney Neoplasms ; metabolism ; pathology ; Male ; Neoplasm Recurrence, Local ; Prognosis ; Racemases and Epimerases ; metabolism

Enzymatic conversion is very important to produce functional rare sugars, but the conversion rate of single enzymes is generally low. To increase the conversion rate, a dual-enzyme coupled reaction system was developed. Dual-enzyme coupled reaction system was constructed using D-psicose-3-epimerase (DPE) and L-rhamnose isomerase (L-RhI), and used to convert D-fructose to D-psicose and D-allose. The ratio of DPE and L-RhI was 1:10 (W/W), and the concentration of DPE was 0.05 mg/mL. The optimum temperature was 60 degrees C and pH was 9.0. When the concentration of D-fructose was 2%, the reaction reached its equilibrium after 10 h, and the yield of D-psicose and D-allose was 5.12 and 2.04 g/L, respectively. Using the dual-enzymes coupled system developed in the current study, we could obtain sugar syrup containing functional rare sugar from fructose-rich raw material, such as high fructose corn syrup.
Aldose-Ketose Isomerases ; metabolism ; Carbohydrate Epimerases ; metabolism ; Fructose ; chemistry ; Glucose ; chemistry ; Hydrogen-Ion Concentration ; Temperature