Surface Display of Designer Protein Scaffolds on Genome-Reduced Strains of Pseudomonas putida
Language English Country United States Media print-electronic
Document type Journal Article, Research Support, Non-U.S. Gov't
- Keywords
- Pseudomonas putida, cellulosome, designer scaffoldin, surface display, synthetic biology,
- MeSH
- beta-Glucosidase metabolism MeSH
- Cellulose metabolism MeSH
- Cellulosomes metabolism MeSH
- Chromosomal Proteins, Non-Histone chemistry MeSH
- Escherichia coli metabolism MeSH
- Genome, Bacterial * MeSH
- Cohesins MeSH
- Membrane Proteins metabolism MeSH
- Metabolic Engineering methods MeSH
- Protein Domains MeSH
- Cell Cycle Proteins chemistry MeSH
- Escherichia coli Proteins metabolism MeSH
- Pseudomonas putida genetics metabolism MeSH
- Recombinant Proteins metabolism MeSH
- Bacterial Outer Membrane metabolism MeSH
- Green Fluorescent Proteins metabolism MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- beta-Glucosidase MeSH
- Cellulose MeSH
- Chromosomal Proteins, Non-Histone MeSH
- Membrane Proteins MeSH
- Cell Cycle Proteins MeSH
- Escherichia coli Proteins MeSH
- Recombinant Proteins MeSH
- Green Fluorescent Proteins MeSH
The bacterium Pseudomonas putida KT2440 is gaining considerable interest as a microbial platform for biotechnological valorization of polymeric organic materials, such as lignocellulosic residues or plastics. However, P. putida on its own cannot make much use of such complex substrates, mainly because it lacks an efficient extracellular depolymerizing apparatus. We seek to address this limitation by adopting a recombinant cellulosome strategy for this host. In this work, we report an essential step in this endeavor-a display of designer enzyme-anchoring protein "scaffoldins", encompassing cohesin binding domains from divergent cellulolytic bacterial species on the P. putida surface. Two P. putida chassis strains, EM42 and EM371, with streamlined genomes and differences in the composition of the outer membrane were employed in this study. Scaffoldin variants were optimally delivered to their surface with one of four tested autotransporter systems (Ag43 from Escherichia coli), and the efficient display was confirmed by extracellular attachment of chimeric β-glucosidase and fluorescent proteins. Our results not only highlight the value of cell surface engineering for presentation of recombinant proteins on the envelope of Gram-negative bacteria but also pave the way toward designer cellulosome strategies tailored for P. putida.
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