Maertens, Barbara and Spriestersbach, Anne and von Groll, Uritza and Roth, Udo and Kubicek, Jan and Gerrits, Michael and Graf, Marcus and Liss, Michael and Daubert, Daniela and Wagner, Ralf and Schaefer, Frank (2010) Gene optimization mechanisms: A multi-gene study reveals a high success rate of full-length human proteins expressed in Escherichia coli. PROTEIN SCIENCE, 19 (7). pp. 1312-1326. ISSN 0961-8368, 1469-896X
Full text not available from this repository. (Request a copy)Abstract
The genetic code is universal, but recombinant protein expression in heterologous systems is often hampered by divergent codon usage. Here, we demonstrate that reprogramming by standardized multi-parameter gene optimization software and de novo gene synthesis is a suitable general strategy to improve heterologous protein expression. This study compares expression levels of 94 full-length human wt and sequence-optimized genes coding for pharmaceutically important proteins such as kinases and membrane proteins in E. coli. Fluorescence-based quantification revealed increased protein yields for 70% of in vivo expressed optimized genes compared to the wt DNA sequences and also resulted in increased amounts of protein that can be purified. The improvement in transgene expression correlated with higher mRNA levels in our analyzed examples. In all cases tested, expression levels using wt genes in tRNA-supplemented bacterial strains were outperformed by optimized genes expressed in non-supplemented host cells.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | CODONS IMMEDIATELY DOWNSTREAM; HIGH-LEVEL EXPRESSION; CELL-FREE SYSTEM; RECOMBINANT PROTEINS; INITIATION CODON; AT-CONTENT; DESIGN; DNA; synthetic genes; gene optimization; heterologous protein expression; fluorescence-based quantification; codon usage; recombinant proteins |
| Subjects: | 600 Technology > 610 Medical sciences Medicine |
| Divisions: | Medicine > Lehrstuhl für Medizinische Mikrobiologie und Hygiene |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 27 Jul 2020 12:52 |
| Last Modified: | 27 Jul 2020 12:52 |
| URI: | https://pred.uni-regensburg.de/id/eprint/24532 |
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