Yuan-Zhi Tan,Liang-Zhi Qiao,Shan-Shan Wang,Juan Zhang,Jing Qian,Min Zhu,Shan-Jing Yao,Dong-Qiang Lin
The enormous potential of mRNA-based therapeutics in vaccines and medicine has led to the consideration of various mRNA sizes, ranging from ∼1000 to 10,000 nucleotides (nt). However, the commonly used Oligo-deoxythymidine (Oligo dT) affinity resins are not initially designed for such a wide range of sizes, and present challenges in achieving optimal performance for the separation of a given mRNA. In this paper, two factors of the resin beads, pore size and grafting were focused. The effects of varying pore sizes, ranging from 150 to 850 nm, on both static and dynamic adsorption for varying-length mRNA (1000–8500 nt) were investigated. It was found that static adsorption capacity and uptake kinetics were correlated with the mRNA length. Interestingly, dynamic binding capacities (DBCs) showed an increase-then-decrease trend with increasing pore size. An optimal pore size of about 350 nm could lead to the highest DBCs for all tested mRNA. Additionally, the effects of polymer grafting on the adsorption performance were studied. The results revealed that polymer-grafted resin exhibited 24–55% higher DBCs than non-grafted resin. The enhanced DBCs could be attributed to the improved uptake rate facilitated by the polymer grafted. These investigations emphasize the importance of optimizing pore size and grafting for Oligo dT affinity resins, providing valuable guidance for selecting and designing new resins for different mRNA products.
Shangyong Li,Linna Wang,Xuehong Chen,Mi Sun,Yantao Han
Chitooligosaccharides (CHOS) have gained increasing attention because of their important biological activities. Enhancing the efficiency of CHOS production essentially requires screening of novel chitosanase with unique characteristics. Therefore, a rapid and efficient one-step affinity purification procedure plays important roles in screening native chitosanases. In this study, we report the design and synthesis of affinity resin for efficient purification of native chitosanases without any tags, using chitodisaccharides (CHDS) as an affinity ligand, to couple with Sepharose 6B via a spacer, cyanuric chloride. Based on the CHDS-modified affinity resin, a one-step affinity purification method was developed and optimized, and then applied to purify three typical glycoside hydrolase (GH) families: 46, 75, and 80 chitosanase. The three purified chitosanases were homogeneous with purities of greater than 95% and bioactivity recovery of more than 40%. Moreover, we also developed a rapid and efficient affinity purification procedure, in which tag-free chitosanase could be directly purified from supernatant of bacterial culture. The purified chitosanases samples using such a procedure had apparent homogeneity, with more than 90% purity and 10–50% yield. The novel purification methods established in this work can be applied to purify native chitosanases in various scales, such as laboratory and industrial scales.Keywords:chitosanases;adsorption analysis;affinity purification
Baoqian Liu,Jianfei Liu,Dongdong Huang,Dong Pei,Jianteng Wei,Duolong Di
Oleuropein is an important natural compound with many biological activities. How to get high purity oleuropein is an urgent issue to be solved. This study provided a method for separating and purifying high purity oleuropein from olive leaves using boric acid affinity resin and a novel solvent system. And the experimental kinetic and adsorption isotherm data of boric acid affinity resin were well–fitted pseudo–first–order kinetic model and Langmuir isotherm model, respectively. The dynamic desorption experiments showed that ethanol was the most efficient eluent while had the lowest oleuropein purity. However, after the novel solvent system treated only one cycle of adsorption and desorption , the purity of oleuropein increased by 5.55 folds from 12.31 ± 0.42% – 68.30 ± 2.08 %. The results can guide the separation and purification of high purity catechol natural products using boric acid affinity resin and mixed solvent systems.
危险品化学品经营许可证（带存储）