earticle

영어

Streptomyces peucetius ATCC 27952 contains two similar genes (1068 nucleotides), dnmL and rmbA, which encode for glucose-1-phosphate (G-1-P) thymidylyltransferases (69% identity and 86% similarity). When the genes were expressed in E. coli under similar conditions, rmbA was expressed in soluble form, and it showed G-1-P thymidylyltransferase activity. But, dnmL was expressed as inclusion bodies. Thorough comparative analysis of DnmL and RmbA by sequence alignment, hydrophobicity scale and homology modeling inferred that the expression of RmbA in soluble form is contributed to the helical structure formed by the high hydrophobicity content in C-terminal sequence of RmbA. Similarly, the formation of inclusion bodies of DnmL was attributed to a loop conformation attained by its C-terminal sequence. To test the significance of these sequences, the C-terminal sequences of DnmL was substituted with that of RmbA. The fusion DnmL was expressed in soluble form and it had similar G-1-P thymidylyltransferase activity as compared to RmbA. These observations demonstrated that the hydrophobic effect plays a significant role in protein folding or structure, and the solubility of the expressed proteins depends on the protein conformations. In conclusion, this method of protein engineering will be a rational tool for enhancing solubility of proteins expressed in E. coli.