Preformulation Characterization and the Effect of Ionic Excipients on the Stability of a Novel DB Fusion Protein.
Preformulation Characterization and the Effect of Ionic Excipients on the Stability of a Novel DB Fusion Protein.
复制标题
DOI：
10.1016/j.xphs.2020.09.008
复制
发表时间：
2021-01
影响因子：
3.8
通讯作者：
Middaugh CR
中科院分区：
文献类型：
--
作者：
Jain A;Hu G;Kumar Ratnakaram SS;Johnson DK;Picking WD;Picking WL;Middaugh CR
志贺氏菌引起的细菌性痢疾在全球儿童和老年人中发病率很高。志贺氏菌的毒力依赖于III型分泌系统（T3SS），该系统将宿主改变效应蛋白注入靶向肠细胞。志贺氏菌T3 SS含有两种组分，侵袭质粒抗原D（IpaD）和侵袭质粒抗原B（IpaB），它们先前被鉴定为广泛保护性抗原。当IpaD和IpaB共表达以得到DB融合（DBF）蛋白时，疫苗效力进一步提高。在各种pH条件下的生物物理表征表明DBF在pH 7和8下最稳定，并且分别在48和50 ° C下失去其构象完整性。强制降解研究表明，DBF的二级结构，三级结构和构象稳定性的显着影响。在磷酸盐缓冲液以及其他阴离子赋形剂的存在下，DBF表现出浓度依赖性构象稳定。分子对接揭示了DBF中可能与植酸相互作用的聚阴离子结合位点。这些位点可用于稳定DBF蛋白。这项工作突出了潜在的不稳定和稳定因素，这不仅提高了我们对DBF蛋白的理解，而且有助于未来开发稳定的志贺氏菌疫苗。
Shigella ssp cause bacillary dysentery (shigellosis) which has high global morbidity in young children and the elderly. The virulence of Shigella relies upon a type III secretion system (T3SS) which injects host altering effector proteins into targeted intestinal cells. The Shigella T3SS contains two components, invasion plasmid antigen D (IpaD) and invasion plasmid antigen B (IpaB), that were previously identified as broadly protective antigens. When IpaD and IpaB were co-expressed to give the DB fusion (DBF) protein, vaccine efficacy was further improved. Biophysical characterization under various pH conditions showed that DBF is most stable at pH 7 and 8 and loses its conformational integrity at 48 and 50 °C respectively. Forced degradation studies revealed significant effects on the secondary structure, tertiary structure and conformational stability of DBF. In the presence of phosphate buffers as well as other anionic excipients, DBF demonstrated a concentration dependent conformational stabilization. Molecular docking revealed potential polyanion binding sites in DBF that may interact with phytic acid. These sites can be exploited to stabilize the DBF protein. This work highlights potential destabilizing and stabilizing factors, which not only improves our understanding of the DBF protein but helps in future development of a stable Shigella vaccine.
登录
查看更多内容
DOI：
10.4049/jimmunol.1302743
发表时间：
2014-02-15
期刊：
Journal of immunology (Baltimore, Md. : 1950)
影响因子：
--
作者：
Heine SJ;Diaz-McNair J;Andar AU;Drachenberg CB;van de Verg L;Walker R;Picking WL;Pasetti MF
通讯作者：
Pasetti MF
影响因子：
2.9
作者：
Jacobson, MP;Pincus, DL;Friesner, RA
通讯作者：
Friesner, RA
影响因子：
3.8
作者：
Wang, Tingting;Kumru, Ozan S.;Volkin, David B.
通讯作者：
Volkin, David B.
影响因子：
2.9
作者：
Barta, Michael L.;Guragain, Manita;Adam, Philip;Dickenson, Nicholas E.;Patil, Mrinalini;Geisbrecht, Brian V.;Picking, Wendy L.;Picking, William D.
通讯作者：
Picking, William D.
影响因子：
8
作者：
Bye, Jordan W.;Falconer, Robert J.
通讯作者：
Falconer, Robert J.