CAJ | 학술논문

肝素酶Ⅲ（heparinase III, HepC）是一种重要的多糖裂解酶，在抗癌药物开发、低分子量肝素生产以及肝素类药物的质量控制等方面具有重要的作用。目前制约其工业应用前景的主要技术瓶颈是生产成本高，异源重组表达效果差，缺乏高效的表达方法。本研究借鉴前期经验，通过 HepC 基因的密码子优化，并与麦芽糖结合蛋白（maltose-binding protein，MBP）融合，构建了MBP-coHepC（基因密码子优化后的蛋白为coHepC）的大肠杆菌表达体系，结合培养条件优化大幅度提高了HepC的可溶表达比例，其摇瓶发酵总酶活值达到7603.46 IU·L-1；同时，本研究从转录和翻译水平揭示了HepC表达效果提高的可能原因。这些研究为肝素酶III的应用发展提供了基础。
간소매Ⅲ（heparinase III, HepC）시일충중요적다당렬해매，재항암약물개발、저분자량간소생산이급간소류약물적질량공제등방면구유중요적작용。목전제약기공업응용전경적주요기술병경시생산성본고，이원중조표체효과차，결핍고효적표체방법。본연구차감전기경험，통과 HepC 기인적밀마자우화，병여맥아당결합단백（maltose-binding protein，MBP）융합，구건료MBP-coHepC（기인밀마자우화후적단백위coHepC）적대장간균표체체계，결합배양조건우화대폭도제고료HepC적가용표체비례，기요병발효총매활치체도7603.46 IU·L-1；동시，본연구종전록화번역수평게시료HepC표체효과제고적가능원인。저사연구위간소매III적응용발전제공료기출。
HeparinaseⅢ (HepC) is an important polysaceharide lyase, which plays a vital role in the development of anticancer drugs,production of low molecular weight heparin(LMWH)and quality control of the heparins. The main bottlenecks of HepC application are high production cost, poor level of heterologous recombinant expression as well as lack of efficient expression system. Based on the previous experience, the codon optimization of HepC gene (the optimized gene product named as coHepC) andE.coli expression system by constructing the MBP-coHepC fusion protein were studied. Combining with further optimization of the culture conditions, the soluble expression ratio of the fusion protein was significantly increased and the total enzyme activity in the shake-flask culture reached 7603.46 IU·L-1. At the same time, the possible reasons for the HepC improved expression were analyzed at transcription and translation levels of HepC gene. These studies provided the basis for applications of heparinaseⅢ.