Abstract
At least 7 proteorhodopsin sequences of Oxyrrhis marina were recently proven in bands obtained by sucrose density gradient centrifugation, and MS analyses revealed that the bands consisted almost of pure, native proteorhodopsins (Rhiel et al. 2020). The proteorhodopsin fractions, i.e., bands B2, B3, and B4 were subjected to transmission electron microscopy. Negative staining revealed that band B2 consisted most likely of monomeric/oligomeric proteorhodopsins with particle dimensions of about 6 nm. Negative staining, freeze-fracture, and cryo-transmission electron microscopy revealed that bands B3 and B4 consisted of vesicular, sheet-like, and cup-shaped structures which all seemed to be composed of protein. Frequently, ring-like protein aggregates were registered at higher magnifications. They measured about 4 nm in diameter with a tiny hole of 1.5 nm in the middle. The bands B2, B3, and B4 were pooled and used to raise an antiserum. Immunoelectron microscopy resulted in intense labeling of the isolated structures. Immunofluorescence light microscopy of formaldehyde-fixed Oxyrrhis cells resulted in intense labeling of the cell periphery. Some cell internal structures became labeled, too. Immunoelectron microscopy of freeze-fractured cells revealed that most likely the membranes of the amphiesmal vesicles were labeled at the cell periphery, while the cell internal label seemed to originate from the food vacuoles.