Influence of the expression vector and its elements on recombinant human prolactin synthesis in Escherichia coli; co-directional orientation of replication and transcription is highly critical.

Autor: Affonso R; Biotechnology Center, Instituto de Pesquisas Energéticas e Nucleares, IPEN - CNEN/SP, São Paulo, SP, Brazil. Electronic address: reginaffonso@yahoo.com.br., Suzuki MF; Biotechnology Center, Instituto de Pesquisas Energéticas e Nucleares, IPEN - CNEN/SP, São Paulo, SP, Brazil. Electronic address: mfsuzuki@ipen.br., Magalhães GS; Immunopathology Laboratory, Butantan Institute, São Paulo, SP, Brazil. Electronic address: geraldo.magalhaes@butantan.gov.br., Bartolini P; Biotechnology Center, Instituto de Pesquisas Energéticas e Nucleares, IPEN - CNEN/SP, São Paulo, SP, Brazil. Electronic address: pbartoli@ipen.br.
Jazyk: angličtina
Zdroj: Journal of microbiological methods [J Microbiol Methods] 2021 Dec; Vol. 191, pp. 106340. Date of Electronic Publication: 2021 Oct 27.
DOI: 10.1016/j.mimet.2021.106340
Abstrakt: The aim of the present work was to define a bacterial expression system that is particularly efficient for the synthesis of recombinant human prolactin (hPRL). In previous work, based on experiments that were basically carried out in parallel with the present ones, the synthesis of rec-hPRL by the p1813-hPRL vector in E. coli HB2151 was >500 mg/L, while it was much lower here (2.5-4-fold), in the RB791 and RRI strains. The highest positive influence on rec-hPRL synthesis was due to the transcription-replication co-orientation of hPRL cDNA and the ori/antibiotic resistance gene, responsible for up to a ~ 5-6-fold higher expression yield. In conclusion, this work confirmed that each bacterial strain of E. coli has a genetic background that can allow a different level of heterologous protein synthesis. The individual study of each element indicated that its action critically depends on the reading orientation in which it is located inside the vector: co-directional orientation of replication and transcription, in fact, greatly increased the level of rec-hPRL expression.
(Copyright © 2021 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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