Comparison of the physical characteristics of chlorosomes from three different phyla of green phototrophic bacteria

• PeakForce Tapping atomic force microscopy was used to image chlorosomes.
• The chlorosomes were from Cfx. aurantiacus, Cba. tepidum and Ca. Cab. thermophilum.
• Nanomechanical AFM was used to assess chlorosome stability.
• We observed heterogeneous fluorescence emission for individual chlorosomes.
• We obtained an AFM image of a chlorosome attached to its cytoplasmic membrane.

Chlorosomes, the major antenna complexes in green sulphur bacteria, filamentous anoxygenic phototrophs, and phototrophic acidobacteria, are attached to the cytoplasmic side of the inner cell membrane and contain thousands of bacteriochlorophyll (BChl) molecules that harvest light and channel the energy to membrane-bound reaction centres. Chlorosomes from phototrophs representing three different phyla, Chloroflexus (Cfx.) aurantiacus, Chlorobaculum (Cba.) tepidum and the newly discovered “Candidatus (Ca.) Chloracidobacterium (Cab.) thermophilum” were analysed using PeakForce Tapping atomic force microscopy (PFT-AFM). Gentle PFT-AFM imaging in buffered solutions that maintained the chlorosomes in a near-native state revealed ellipsoids of variable size, with surface bumps and undulations that differ between individual chlorosomes. Cba. tepidum chlorosomes were the largest (133 × 57 × 36 nm; 141,000 nm3 volume), compared with chlorosomes from Cfx. aurantiacus (120 × 44 × 30 nm; 84,000 nm3) and Ca. Cab. thermophilum (99 × 40 × 31 nm; 65,000 nm3). Reflecting the contributions of thousands of pigment–pigment stacking interactions to the stability of these supramolecular assemblies, analysis by nanomechanical mapping shows that chlorosomes are highly stable and that their integrity is disrupted only by very strong forces of 1000–2000 pN. AFM topographs of Ca. Cab. thermophilum chlorosomes that had retained their attachment to the cytoplasmic membrane showed that this membrane dynamically changes shape and is composed of protrusions of up to 30 nm wide and 6 nm above the mica support, possibly representing different protein domains. Spectral imaging revealed significant heterogeneity in the fluorescence emission of individual chlorosomes, likely reflecting the variations in BChl c homolog composition and internal arrangements of the stacked BChls within each chlorosome.