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KMID : 0578320100300020127
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Molecules and Cells
2010 Volume.30 No. 2 p.127 ~ p.135
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N-Terminal pI Determines the Solubility of a Recombinant Protein Lacking an Internal Transmembrane-like Domain in E. coli
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Lee Sang-Jun
Han Yun-Hee
Kim Young-Ok
Nam Bo-Hye
Kong Hee-Jeong
Kim Kyung-Kil
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Abstract
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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.
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KEYWORD
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ΔGRNA value, feedback regulatory mechanism, hydrophilicity, inner membrane channel, pI value trigger (facilitated diffusion)
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