The amino acid sequences in the C-terminal region of glucose-1-phosphate thymidylyltransferases determine their soluble expression in Escherichia coli

Protein Engineering, Design & Selection : PEDS
Bijay SinghJae Kyung Sohng

Abstract

Two similar genes, dnmL and rmbA in Streptomyces peucetius, which encode for glucose-1-phosphate (G-1-P) thymidylyltransferases were expressed in Escherichia coli under similar conditions. While RmbA was expressed in soluble form, DnmL was found as insoluble aggregates in inclusion bodies. The difference in expression of these similar proteins led to investigate into the amino acid sequences of these proteins by sequence alignment, hydrophobicity scale and homology modeling. These analyses showed that the two proteins are different only in the C-terminal sequences. Deletion of C-terminal sequence of DnmL increased the expression level of truncated DnmL. Substitution of C-terminal sequence of DnmL with RmbA also expressed the recombinant protein in soluble form. Finally, mutation of six amino acids in DnmL rendered the protein expressed in soluble form. These results suggested that the soluble expression of the thymidylyltransferases lies in the C-terminal sequences. In conclusion, these methods of protein engineering will be a rational tool for enhancing solubility of proteins expressed in E.coli.

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Citations

Aug 16, 2012·Applied Microbiology and Biotechnology·Bijay SinghJae Kyung Sohng
Apr 1, 2018·Applied Microbiology and Biotechnology·Nguyen Huy ThuanJae Kyung Sohng

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