Li Yang-Nan, Zhu Chen-Yuan, Xu Chen-Lu, Yu Shen, Huan Tong, Zhang Ye-Wang
School of Pharmacy, Jiangsu University, Zhenjiang, 212013, PR China.
World J Microbiol Biotechnol. 2024 Dec 23;41(1):15. doi: 10.1007/s11274-024-04225-2.
A heparinase III (NsHep-III) from Niabella sp. was identified, cloned, and expressed as soluble form in E. coli BL21 (DE3). With heparin as substrate, the maximum activity of NsHep-III of 90.0 U·mg was achieved at pH 7.1 Tris-HCl (containing 15 mM Mg) and 30C. The half-life of NsHep-III was determined to be 5 h at 30C. The interactions between NsHep-III and substrates were studied by molecular docking. The combination of NsHep-III and a heparinase I from Bacteroides eggthii (BeHep-I) was employed to cleave heparin. Analysis of the enzymatic products of NsHep-III by SAX-HPLC showed seven different modified disaccharides, indicating that NsHep-III has a wide range of substrate specificity. The results of GPC analysis demonstrated that the average molecular weight of the product of heparin cleavage by the combination of NsHep-III/BeHep-I was reduced to 3969 Da, which accounted for 90% of all the components, and complied with the requirements of the European Pharmacopoeia. NsHep-III has notable activity and efficiency in cleaving heparin, which is potentially useful for the industrial production of low molecular weight heparin.
从Niabella sp.中鉴定、克隆出一种肝素酶III（NsHep-III），并在大肠杆菌BL21（DE3）中以可溶性形式表达。以肝素为底物时，NsHep-III在pH 7.1的Tris-HCl（含15 mM Mg）和30℃条件下，最大活性达到90.0 U·mg。NsHep-III在30℃时的半衰期测定为5小时。通过分子对接研究了NsHep-III与底物之间的相互作用。将NsHep-III与来自多形拟杆菌的肝素酶I（BeHep-I）联合用于裂解肝素。通过SAX-HPLC对NsHep-III的酶解产物分析显示有七种不同的修饰二糖，表明NsHep-III具有广泛的底物特异性。凝胶渗透色谱（GPC）分析结果表明，NsHep-III/BeHep-I联合裂解肝素产物的平均分子量降至3969 Da，占所有组分的90%，符合欧洲药典的要求。NsHep-III在裂解肝素方面具有显著的活性和效率，这对低分子量肝素的工业化生产具有潜在用途。
