期刊
JOURNAL OF BIOTECHNOLOGY
卷 120, 期 1, 页码 11-24出版社
ELSEVIER
DOI: 10.1016/j.jbiotec.2005.05.028
关键词
codon usage; Gillespie algorithm; translation; PDI; recombinant protein; Escherichia coli; computer simulation
Recombinant human protein disulfide isomerase (PDI) was expressed in vivo in Escherichia coli using a non-optimised gene sequence and an optimised sequence with four 5' codons substituted by synonymous codons that take less time to translate. The optimisation resulted in a 2-fold increase of total PDI concentration and by successive optimisation with expression at low temperature in a 10-fold increase of the amount of soluble PDI in comparison with the original wild-type construct. The improvement can be due to a faster clearing of the ribosome binding site on the mRNA, elevating the translation initiation rate and resulting in higher ribosome loading and better ribosome protection of the PDI mRNA against endonucleolytic cleavage by RNase. This hypothesis was supported by a novel computer simulation model of E. coli translational ribosome traffic based upon the stochastic Gillespie algorithm. The study indicates the applicability of such models in optimisation of recombinant protein sequences. (c) 2005 Elsevier B.V All rights reserved.
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