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Fig. 4 | BMC Biology

Fig. 4

From: NmPin from the marine thaumarchaeote Nitrosopumilus maritimus is an active membrane associated prolyl isomerase

Fig. 4

Localization of endogenous NmPin in N. maritimus observed by microscopy. a Overlay of brightfield (grey) and fluorescence (blue, DAPI stain) microscopy images (bar: 5 μm). Cells exhibit a normal rod-shaped form after paraformaldehyde (PFA) fixation before harvesting by centrifugation as observed by transmission electron microscopy (TEM) (insert, bar: 500 nm). b Fluorescence microscopy of PFA-fixed N. maritimus (bars: 5 μm). Top left, immuno-staining of endogenous NmPin with Alexa488. Top right, DAPI staining of DNA. Bottom left, overlay of NmPin and DAPI stained cells. The square indicates the area of magnification. Bottom right, magnification of NmPin stained cells show a localization in the cell envelope (bar: 2 μm). c Detailed TEM microscopy images of PFA-fixed N. maritimus cells show a surface layer (S-layer) enframing the cytoplasmic membrane (arrowheads, bar: 50 nm, bar insert: 20 nm). d Overlay of brightfield (grey) and DAPI stain (blue) images of N. maritimus without PFA fixation before harvesting (bar: 5 μm). The insert shows a TEM image confirming a change in cell shape to a more spheroidal form with a very diffuse membrane structure (bar: 100 nm). e Fluorescence microscopy images of N. maritimus without PFA fixation before harvesting (bars: 5 μm). Top left, NmPin fluorescence-staining with Alexa488. Top right, DAPI stain. Bottom left, overlay of NmPin and DAPI stained cells. The square indicates the area of magnification. Bottom right image shows a magnification of the NmPin stain with a less uniform localization in the cell envelope (bar: 2 μm). f TEM microscopy picture of Immunogold-labelled NmPin in N. maritimus without PFA treatment (bars: 100 nm). Black spots representing NmPin are concentrated in the outer diffuse membrane area of N. maritimus cells. g Fluorescence microscopy images of N. maritimus incubated in low salt conditions (phosphate buffered saline; PBS) (bars: 5 μm). Top left, picture of immuno-stained NmPin with Alexa488, showing a significant decrease of fluorescence intensity in comparison to marine salt conditions. Top right, DAPI stained cells. Bottom left, overlay of DAPI and NmPin stained cells. The square indicates the area of magnification. Bottom right, the magnification of NmPin stain confirms the signal intensity loss (bar: 2 μm). h TEM microscopy image of Immunogold-labelled NmPin in N. maritimus after incubation in PBS (bar: 100 nm). Black spots representing NmPin are localized arbitrarily and show a detachment of NmPin from the cell envelope. i Schematic presentation of NmPin localization in N. maritimus

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