Engineering a suite of E. coli strains for enhanced expression of bacterial polysaccharides and glycoconjugate vaccines.
Engineering a suite of E. coli strains for enhanced expression of bacterial polysaccharides and glycoconjugate vaccines.
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DOI：
10.1186/s12934-022-01792-7
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发表时间：
2022-04-21
影响因子：
6.4
通讯作者：
中科院分区：
文献类型：
--
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在生物技术主力细菌大肠杆菌中，糖工程是一个快速发展的领域，特别是用于生产糖缀合物疫苗候选物（生物缀合）。糖缀合物的有效生产需要在细菌细胞内及时协调表达三种组分：载体蛋白、聚糖抗原和偶联酶。因此，选择合适的E.大肠杆菌宿主细胞是至关重要的。微生物底盘工程长期以来一直用于提高化学品和生物聚合物的产量，但其在疫苗生产中的应用很少。在这项研究中，我们设计了一个家庭的11 E。通过去除和/或添加合理选择的组分来增强肺炎链球菌荚膜多糖的表达，从而提高肺炎球菌结合疫苗的产量。重要的是，所有菌株都表达解毒形式的内毒素，一种在细菌细胞中产生的治疗剂的相关污染物。使用CRISPR以迭代方式改变每个菌株的基因组背景，以产生没有抗生素标记或瘢痕序列的菌株。在本研究中产生的11个改造菌株中，E. coli Falcon、Peregrine和Sparrowwhawk均表现出S.肺炎血清型4胶囊。鹰（一种不含肠细菌共同抗原的菌株，含有GalNAc差向异构酶和从染色体表达的PglB）和雀鹰（不含肠杆菌共同抗原、O-抗原连接酶和链长决定簇，含有GalNAc差向异构酶和来自肺炎链球菌的链长调节剂的菌株）分别产生AcrA-SP 4缀合物，其聚糖比基础菌株中产生的聚糖多4倍和14倍，W3110除了应用于肺炎球菌候选疫苗的生产外，11种新菌株的库将成为糖工程社区的宝贵资源。在线版本包含补充材料，可通过10.1186/s12934-022-01792-7获得。
Glycoengineering, in the biotechnology workhorse bacterium, Escherichia coli, is a rapidly evolving field, particularly for the production of glycoconjugate vaccine candidates (bioconjugation). Efficient production of glycoconjugates requires the coordinated expression within the bacterial cell of three components: a carrier protein, a glycan antigen and a coupling enzyme, in a timely fashion. Thus, the choice of a suitable E. coli host cell is of paramount importance. Microbial chassis engineering has long been used to improve yields of chemicals and biopolymers, but its application to vaccine production is sparse. In this study we have engineered a family of 11 E. coli strains by the removal and/or addition of components rationally selected for enhanced expression of Streptococcus pneumoniae capsular polysaccharides with the scope of increasing yield of pneumococcal conjugate vaccines. Importantly, all strains express a detoxified version of endotoxin, a concerning contaminant of therapeutics produced in bacterial cells. The genomic background of each strain was altered using CRISPR in an iterative fashion to generate strains without antibiotic markers or scar sequences. Amongst the 11 modified strains generated in this study, E. coli Falcon, Peregrine and Sparrowhawk all showed increased production of S. pneumoniae serotype 4 capsule. Eagle (a strain without enterobacterial common antigen, containing a GalNAc epimerase and PglB expressed from the chromosome) and Sparrowhawk (a strain without enterobacterial common antigen, O-antigen ligase and chain length determinant, containing a GalNAc epimerase and chain length regulators from Streptococcus pneumoniae) respectively produced an AcrA-SP4 conjugate with 4 × and 14 × more glycan than that produced in the base strain, W3110. Beyond their application to the production of pneumococcal vaccine candidates, the bank of 11 new strains will be an invaluable resource for the glycoengineering community. The online version contains supplementary material available at 10.1186/s12934-022-01792-7.
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影响因子：
5.8
作者：
Cuccui J;Terra VS;Bossé JT;Naegeli A;Abouelhadid S;Li Y;Lin CW;Vohra P;Tucker AW;Rycroft AN;Maskell DJ;Aebi M;Langford PR;Wren BW;BRaDP1T Consortium
通讯作者：
BRaDP1T Consortium
DOI：
10.1073/pnas.86.7.2172
发表时间：
1989-04-01
影响因子：
11.1
作者：
CHUNG, CT;NIEMELA, SL;MILLER, RH
通讯作者：
MILLER, RH
影响因子：
3.7
作者：
Blank K;Hensel M;Gerlach RG
通讯作者：
Gerlach RG
影响因子：
5.6
作者：
Hagelueken, Gregor;Huang, Hexian;Mainprize, Iain L.;Whitfield, Chris;Naismith, James H.
通讯作者：
Naismith, James H.
影响因子：
4.8
作者：
Bernatchez, S;Szymanski, CM;Wakarchuk, WW
通讯作者：
Wakarchuk, WW