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Agroinfiltration contributes to VP1 recombinant protein degradation
Priyen Pillay1, Karl J Kunert1, Stefan van Wyk1
1a Department of Plant and Soil Sciences , Forestry and Agricultural Biotechnology Institute (FABI), University of Pretoria , Hillcrest, Pretoria , South Africa.
Bioengineered
|July 27, 2016
Summary
Tobacco agroinfiltration for recombinant protein production is hindered by proteases. This study shows recombinant proteins are sensitive to proteases, and agroinfiltration increases protease expression, potentially reducing protein yield.
Area of Science:
- Plant biotechnology
- Molecular biology
- Biochemistry
Background:
- Tobacco agroinfiltration is a promising system for recombinant protein production.
- Protease activity in plants can degrade target proteins, limiting production efficiency.
Purpose of the Study:
- To investigate the protease sensitivity of model recombinant proteins.
- To identify plant proteases induced by agroinfiltration that may affect protein yield.
Main Methods:
- Protease sensitivity assays using trypsin, papain, and plant extracts.
- In silico analysis of protease cleavage sites and protein-protease interactions.
- RNA sequencing (RNA-seq) to identify differentially expressed protease transcripts post-agroinfiltration.
Main Results:
- Recombinant VP1 showed higher sensitivity to trypsin than papain.
- Agroinfiltrated tobacco tissues exhibited elevated cathepsin and legumain activities.
- Three candidate protease genes, including RD21, NbVPE1a, and NbVPE1b, were identified as differentially expressed.
Conclusions:
- Recombinant proteins are susceptible to degradation by plant proteases.
- Agroinfiltration upregulates specific protease genes in tobacco.
- Understanding and managing protease activity is crucial for optimizing recombinant protein production in plants.
