Prokaryotic expression of a hub protein of white spot syndrome virus with vaccine potential

Lu, Yongzhong; Cheng, Linyue; Wang, Fang

doi:10.1111/are.14690

Cited by 2 publications

(2 citation statements)

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“…Prokaryotic protein expression systems are usually the first choice for protein heterologous expression because of their obvious advantages. For example, the envelope protein VP62 of the white spot syndrome virus, a protein with vaccine potential, was expressed and purified in E. coli (Lu et al 2020). The novel antibacterial fusion protein cecropin B-human lysozyme was also expressed in E. coli (Zhang et al 2019).…”

Section: Discussionmentioning

confidence: 99%

“…Some inclusion bodies can become soluble proteins by altering the expression vectors or by changing the culture conditions such as culture temperature, IPTG concentration and induction times. However, other inclusion bodies remain insoluble proteins regardless of alterations to the expression vectors or culture conditions (Makrides 1996;Rabhi-Essafi et al 2017;Lu et al 2020). It is highly likely that the insolubility may be related to the lack of a post-translational modification mechanism and/or the lack of respective chaperon proteins for the target protein in the heterologous prokaryotic expression host E. coli (Sui et al 2018).…”

Section: Discussionmentioning

confidence: 99%

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Prokaryotic expression and solubilisation of Arabidopsis ROOT UVB SENSITIVE 1 from inclusion bodies in Escherichia coli

Hou

Tong

2022

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The RUS (ROOT UVB SENSITIVE 1) proteins characterized by their unique DUF647 domain are widely distributed in eukaryotes. Their functional roles are largely unknown except for the possible involvement of Arabidopsis RUS1 and RUS2 in early seedling development. To investigate the biochemical roles of the RUS proteins, full length and truncated Arabidopsis RUS1 were seamlessly fused with GFP and cloned into prokaryotic expression vector pQE-100 which allows proteins expressed with an N-terminal 6?His tag. Expression of the full length RUS1-GFP could not be detected after adding the inducer IPTG, while a truncated RUS1-GFP was expressed at high levels and formed inclusion bodies in Escherichia coli. The inclusion bodies were dissolved in a denaturing buffer, and then the truncated RUS1-GFP fusion protein in the supernatant was bound to a Ni-NTA slurry. The bound proteins were eluted after the non-specific binding proteins were washed away. The purified truncated proteins were detected as a single clear band of the expected size in SDS-PAGE, and were further confirmed by the Western blot test. Our results suggest that the impossible expression of the full length RUS1 protein in E. coli can be expressed in truncated form, and inclusion bodies can be effectively solubilized.

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Section: Discussionmentioning

confidence: 99%