Article Figures & Data
Figures
- FIG 1
Study overview. (A) Diverse phytases were selected following a search of the Joint Genome Institute’s Integrated Microbial Genomes and Microbiomes (IMG/M) database. Figure includes the three classes of phytase enzymes in microbes: cysteine phytases (CPs), histidine acid phytases (HAPs), and beta-propeller phytases (BPPs). (B) Genes were optimized for expression in Proteobacteria, synthesized, and engineered into three bacterial hosts. (C) Engineered strains were evaluated in vitro for their ability to hydrolyze phytate and release Pi. (D) Arabidopsis was inoculated with the best-performing strains and monitored for improvements in fresh and dry weights and rosette size.
- FIG 2
Phytase activity of engineered strains in liquid culture. Pi levels in culture supernatant where phytate is the only phosphate source at 10 days for each construct expressed in P. putida KT2440, P. simiae WCS417r, and Ralstonia sp. PGPB with engineered phytases are screened for the ability to release Pi from phytate using the malachite green-based QuantiChrom phosphate assay kit (BioAssay Systems). Each category of enzyme is ordered by amino acid length, from shortest to longest. Gray space indicates no measurement taken. *, strains used in plant assays; $, E. coli AppA included as positive control.
- FIG 3
Effect of pH on phytase activity. (A to D) Phytase activity in the extracellular medium for H07 (A), H19 (B), H11 (C), and C24 (D). The values are the averages from three experiments. The tested phytases have optimal activities across a range of pHs.
- FIG 4
Engineered strains improve plant growth under Pi-limited conditions. Arabidopsis was grown on one-half-strength Murashige and Skoog agar supplemented with 0.8 mM sodium phytate and specific bacterial treatments. (A to F) Representative images 17 days postinoculation with one-quarter-strength Ringer’s solution (A), P. simiae WCS417r (B), or P. simiae:H07 (C) or 19 days postinoculation with one-quarter-strength Ringer’s solution (D), Ralstonia sp. 352_2 (E), or Ralstonia sp:H11 (F). (G) Fold change in rosette size, fresh weight, and dry weight upon engineered and unengineered strain treatments versus mock inoculation for all strains tested. Brown bars indicate control strains without engineered phytases. Individual plant photos in B, C, E, and F are colored to correspond with data in G.
Additional Files
Supplemental material
- Supplemental file 1 -
Tables S1 and S4, Fig. S1 to S9, Notes S1 and S2
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- Supplemental file 2 -
Table S2
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- Supplemental file 3 -
Table S3
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- Supplemental file 1 -
