Concomitant use of immobilized uridine-cytidine kinase and polyphosphate kinase for 5'-cytidine monophosphate production / 生物工程学报

Uridine-cytidine kinase, an important catalyst in the compensation pathway of nucleotide metabolism, can catalyze the phosphorylation reaction of cytidine to 5'-cytidine monophosphate (CMP), but the reaction needs NTP as the phosphate donor. To increase the production efficiency of CMP, uridine-cytidine kinase gene from Thermus thermophilus HB8 and polyphosphate kinase gene from Rhodobacter sphaeroides were cloned and expressed in Escherichia coli BL21(DE3). Uridine-cytidine kinase was used for the generation of CMP from cytidine and ATP, and polyphosphate kinase was used for the regeneration of ATP. Then, the D403 metal chelate resin was used to adsorb Ni²⁺ to form an immobilized carrier, and the immobilized carrier was specifically combined with the recombinant enzymes to form the immobilized enzymes. Finally, single-factor optimization experiment was carried out to determine the reaction conditions of the immobilized enzyme. At 30 °C and pH 8.0, 60 mmol/L cytidine and 0.5 mmol/L ATP were used as substrates to achieve 5 batches of high-efficiency continuous catalytic reaction, and the average molar yield of CMP reached 91.2%. The above method has the advantages of low reaction cost, high product yield and high enzyme utilization rate, and has good applied value for industrial production.

Overexpression of Uridine-Cytidine Kinase 2 Correlates with Breast Cancer Progression and Poor Prognosis / 한국유방암학회지

PURPOSE: Uridine-cytidine kinase (UCK) 2 is a rate-limiting enzyme involved in the salvage pathway of pyrimidine-nucleotide biosynthesis. Recent studies have shown that UCK2 is overexpressed in many types of cancer and may play a crucial role in activating antitumor prodrugs in human cancer cells. In the current study, we evaluated the potential prognostic value of UCK2 in breast cancer. METHODS: We searched public databases to explore associations between UCK2 gene expression and clinical parameters in patients with breast cancer. Gene set enrichment analysis (GSEA) was performed to identify biological pathways associated with UCK2 gene expression levels. Survival analyses were performed using 10 independent large-scale breast cancer microarray datasets. RESULTS: We found that UCK2 mRNA expression was elevated in breast cancer tissue compared with adjacent nontumorous tissue or breast tissue from healthy controls. High UCK2 levels were correlated with estrogen receptor negativity (p<0.001), advanced tumor grade (p<0.001), and poor tumor differentiation (p<0.001). GSEA revealed that UCK2-high breast cancers were enriched for gene sets associated with metastasis, progenitor-like phenotypes, and poor prognosis. Multivariable Cox proportional hazards regression analyses of microarray datasets verified that high UCK2 gene expression was associated with poor overall survival in a dose-response manner. The prognostic power of UCK2 was superior to that of TNM staging and comparable to that of multiple gene signatures. CONCLUSION: These findings suggest that UCK2 may be a promising prognostic biomarker for patients with breast cancer.