Article Figures & Data
Figures
- FIG. 1.
Granule formation through micelle formation from hydroxybutyrate chains covalently attached to PhaC. The colored spheres shown on amorphous PHB represent proteins that have been shown to or are proposed to associate with the granule surface. Gray spheres, PhaC; blue spheres, PhaP; red spheres, PhaR; green spheres, PhaZ1a; purple sphere, PhaZ1b; orange sphere, PhaZ1c; brown diamond, oligomer hydrolase. nHBCoA, hydroxybutyryl coenzyme A.
- FIG. 2.
Granule formation through budding from the inner leaflet of the plasma membrane. nHBCoA, hydroxybutyryl coenzyme A.
- FIG. 3.
TEM images of wild-type W. eutropha during growth in PHBP and PHBU. (A) PHBP at 2.5 h; (B) PHBP at 5 h; (C) PHBP at 9 h; (D) PHBP at 24 h; (E) PHBP at 73 h; (F) PHBU at 48 h. All images were recorded at the same magnification (bars, 3 μm). The changes in cell size for the samples are reflected by the histograms below the images. The histograms indicate the size distribution of cell profiles (obtained by measuring the major axis of all cell profiles). Images other than the ones shown were also used for this analysis.
- FIG. 4.
TEM images of wild-type W. eutropha grown in TSB for 24 h (A) and wild-type W. eutropha grown in PHBP for 2.5 h after 24 h in TSB (B). The two images are the same magnification (bars, 4.5 μm).
- FIG. 5.
TEM images of wild-type W. eutropha at 2.5 h in PHBP. Nascent granules are localized on dark-stained mediation elements near the center of the cell. Bars, 0.5 μm. (A) The arrow indicates a dark-stained mediation element. (B) Cell at higher magnification.
- FIG. 6.
TEM images of wild-type W. eutropha in PHBP at 2.5 h (A), 5 h (B), 9 h (C), and 24 h (D), showing the “disappearance” of the dark-stained mediation elements. Bar, 0.5 μm.
- FIG. 7.
TEM image of W. eutropha at 73 h in PHBP. Significant granule coalescence (black arrows) was observed. In addition, small granules of unknown composition (white arrow) are present. Bar, 0.8 μm.
- FIG. 8.
TEM image of W. eutropha at 48 h in PHBU. There seemed to be different populations of cells. Some cells contained a high number of smaller granules (compared to those at 24 h in PHBP), while others contained a few large granules. Bar, 0.5 μm.
- FIG. 9.
TEM images of consecutive serial sections (A→B→C) of the sample at 5 h in PHBP. The longest cell profile appears to result from a cell that is lying parallel to the plane of the sectioning knife, since its position does not change relative to the other two cell profiles (arrows), which seem to result from cells that were lying perpendicular to the place of section. Since the length of the long cell profile does not change from panel A to panel B, this length represents the actual length of the cell. Notice that the cell length in panel C is less. These images also reveal that the size of the granules is fairly uniform. Bar, 1 μm.
Tables
- TABLE 1.
Estimated average cell volume and total surface area of granules per W. eutropha H16 cell determined by the stereology method and reported as a function of time and cultivation conditionsa
Sample ACP/NCP (μm2)b CE (%)c VC (μm3)d SG (μm2)e TSB, 4 hf 0.84 8 0.9-1.8 1.3-2.5 TSB, 24 h 0.42 6 0.9 1.3 PHBP, 2.5 hf 1.09 6 1.2-2.3 NDg PHBP, 5 hf 0.82 5 0.9-1.8 NDg PHBP, 9 h 0.50 10 1.1 3.6 PHBP, 24 h 0.55 8 1.2 7.3 PHBP, 73 h 0.64 9 1.4 11.2h PHBU, 48 h 0.49 4 NDi NDi ↵ a The change in cell volume is also reflected in the ACP/NCP values. Note that the coefficient of error associated with ACP/NCP is also the minimum coefficient of error associated with VC.
↵ b Total area of cell profiles/number of cell profiles.
↵ c CE, coefficient of error. The sampling error associated with ACP/NCP also applies to VC.
↵ d Average volume of a cell.
↵ e Average total surface area of granules per cell.
↵ f Cells are dividing at these times; ranges of VC and SG are shown.
↵ g ND, not determined since granules were too small and ill defined.
↵ h The tiny granules observed at this time were also counted during measurement.
↵ i ND, not determined since there seemed to be different phenotypes of cells.
- TABLE 2.
Estimated average cell volumes calculated using the average length of the longest cell profiles (∼4% of the total cell profiles measured) at each time and average widths of all cell profiles in images used to measure the length of the cell profilesa
Sample No. of cell profiles measured Width (μm)b Avg cell vol (μm3) PHBP, 2.5 h 482 0.75 ± 0.015 2.4c PHBP, 5 h 768 0.69 ± 0.014 1.7c PHBP, 9 h 1,596 0.66 ± 0.007 1.2 PHBP, 24 h 600 0.68 ± 0.011 1.3 PHBP, 73 h 754 0.74 ± 0.011 1.5
