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Mol. Cells 2010; 30(2): 127-135

Published online July 23, 2010

https://doi.org/10.1007/s10059-010-0097-9

© The Korean Society for Molecular and Cellular Biology

N-Terminal pI Determines the Solubility of a Recombinant Protein Lacking an Internal Transmembrane-like Domain in E. coli

Sang Jun Lee*, Yun Hee Han, Young Ok Kim, Bo Hye Nam, Hee Jeong Kong, and Kyung Kil Kim

Biotechnology Research Division, National Fisheries Research and Development Institute, Pusan 619-902, Korea

Correspondence to : *Correspondence: sangjl@nrfdi.go.kr

Received: February 10, 2010; Revised: April 16, 2010; Accepted: April 26, 2010

Abstract

We examined whether the isoelectric point (pI) of the N-terminal region of the recombinant protein 7xMefp1 acts as a universal index for expression of the protein in solu-ble form in E. coli. Expression analysis of 7xMefp1 fused to various N-terminal sequences with pI values ranging from 2.73 to 13.35 yielded three pI range-specific curves (acidic, neutral, and alkaline curves at pI 2.73-3.25, 4.61-9.58, and 9.90-13.35, respectively) for soluble expression (by facilitated diffusion) as a proportion of total protein. For neutral N-termini (pI 4.61-9.58), the total amount of rMefp1 expressed was minimally affected by ΔGRNA for unfolding the mRNA secondary structure. The highly hydrophilic nature of longer N-terminal sequences with strongly acidic and alkaline pI values reduced the transla-tion of rMefp1-encoding transcripts, thereby reducing the amount of soluble rMefp1 produced. After characterizing both feedback and non-feedback regu-lation in the acidic, alkaline, and neutral pI ranges, we suggest that three different pI range-specific soluble expression curves exist for the recombinant protein, each defined by specific ranges of the leader sequence pI values.

Keywords ΔGRNA value, feedback regulatory mechanism, hydrophilicity, inner membrane channel, pI value trigger (facilitated diffusion)

Article

Research Article

Mol. Cells 2010; 30(2): 127-135

Published online August 31, 2010 https://doi.org/10.1007/s10059-010-0097-9

Copyright © The Korean Society for Molecular and Cellular Biology.

N-Terminal pI Determines the Solubility of a Recombinant Protein Lacking an Internal Transmembrane-like Domain in E. coli

Sang Jun Lee*, Yun Hee Han, Young Ok Kim, Bo Hye Nam, Hee Jeong Kong, and Kyung Kil Kim

Biotechnology Research Division, National Fisheries Research and Development Institute, Pusan 619-902, Korea

Correspondence to:*Correspondence: sangjl@nrfdi.go.kr

Received: February 10, 2010; Revised: April 16, 2010; Accepted: April 26, 2010

Abstract

We examined whether the isoelectric point (pI) of the N-terminal region of the recombinant protein 7xMefp1 acts as a universal index for expression of the protein in solu-ble form in E. coli. Expression analysis of 7xMefp1 fused to various N-terminal sequences with pI values ranging from 2.73 to 13.35 yielded three pI range-specific curves (acidic, neutral, and alkaline curves at pI 2.73-3.25, 4.61-9.58, and 9.90-13.35, respectively) for soluble expression (by facilitated diffusion) as a proportion of total protein. For neutral N-termini (pI 4.61-9.58), the total amount of rMefp1 expressed was minimally affected by ΔGRNA for unfolding the mRNA secondary structure. The highly hydrophilic nature of longer N-terminal sequences with strongly acidic and alkaline pI values reduced the transla-tion of rMefp1-encoding transcripts, thereby reducing the amount of soluble rMefp1 produced. After characterizing both feedback and non-feedback regu-lation in the acidic, alkaline, and neutral pI ranges, we suggest that three different pI range-specific soluble expression curves exist for the recombinant protein, each defined by specific ranges of the leader sequence pI values.

Keywords: ΔGRNA value, feedback regulatory mechanism, hydrophilicity, inner membrane channel, pI value trigger (facilitated diffusion)

Mol. Cells
Mar 31, 2023 Vol.46 No.3, pp. 131~189
COVER PICTURE
The physiologically important cytoprotective signaling in normal cells (background area in turquoise) mediated by NRF2 (blue chain) is often hijacked by cancer cells (red ball) in the tumor microenvironment (yellow area). However, the differential roles of NRF2 throughout the multistage carcinogenesis remains largely unresolved (white-colored overlapping misty areas).

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