期刊
FRONTIERS IN PLANT SCIENCE
卷 11, 期 -, 页码 -出版社
FRONTIERS MEDIA SA
DOI: 10.3389/fpls.2020.00868
关键词
mass spectrometry; proteotypic peptide; QconCAT; Synthetic Biology; Modular Cloning; photosynthesis; Chlamydomonas reinhardtii
资金
- Deutsche Forschungsgemeinschaft [TRR 175]
- Landesforschungsschwerpunkt BioComp
The productivity of plants and microalgae needs to be increased to feed the growing world population and to promote the development of a low-carbon economy. This goal can be achieved by improving photosynthesis via genetic engineering. In this study, we have employed the Modular Cloning strategy to overexpress the Calvin-Benson cycle (CBC) enzyme sedoheptulose-1,7-bisphosphatase (SBP1) up to threefold in the unicellular green algaChlamydomonas reinhardtii. The protein derived from the nuclear transgene represented similar to 0.3% of total cell protein. Photosynthetic rate and growth were significantly increased in SBP1-overexpressing lines under high-light and elevated CO(2)conditions. Absolute quantification of the abundance of all other CBC enzymes by the QconCAT approach revealed no consistent differences between SBP1-overexpressing lines and the recipient strain. This analysis also revealed that the 11 CBC enzymes represent 11.9% of total cell protein inChlamydomonas. Here, the range of concentrations of CBC enzymes turned out to be much larger than estimated earlier, with a 128-fold difference between the most abundant CBC protein (rbcL) and the least abundant (triose phosphate isomerase). Accordingly, the concentrations of the CBC intermediates are often but not always higher than the binding site concentrations of the enzymes for which they act as substrates. The enzymes with highest substrate to binding site ratios might represent good candidates for overexpression in subsequent engineering steps.
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