Open AccessThe structure difference of proteins isolated on substrate with different techniques as studied by the atomic force microscope
Author(s) -
Umemura K.,
Sutoh K.,
Tokunaga F.,
Kataoka M.,
Kamkubo H.,
Arakawa H.,
Ikai A.
Publication year - 1996
Publication title -
scanning
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.359
H-Index - 47
eISSN - 1932-8745
pISSN - 0161-0457
DOI - 10.1002/sca.1996.4950180403
Subject(s) - mica , calibration curve , substrate (aquarium) , chemistry , protein filament , atomic force microscopy , molecule , deposition (geology) , analytical chemistry (journal) , crystallography , materials science , nanotechnology , chromatography , composite material , paleontology , biochemistry , oceanography , organic chemistry , sediment , detection limit , biology , geology
The height and width of proteins deposited on mica substrates were measured from cross sections of their atomic force microscope (AFM) images. The tapping mode AFM gave very stable and reproducible images for high molecular weight proteins. The following results were obtained: (1) The thickness of mono‐, bi‐, and trilayered purple membranes was 5.3, 10.4, and 16.0 nm, respectively. The calibration curve of z range of AFM based on the above data was linear. The deviation of the calibration curve at the origin was < 0.6 nm. (2) The height of slow form α‐2‐macroglobulin (α2M) molecule changed depending on sampling methods. When the protein was freeze‐dried on a mica substrate prewetted with water, α2M gave the highest value for its height. The fact that freeze‐drying, especially after prewetting of the substrate, was effective to prevent flattening of the molecule suggested that sample deposition must be as gentle as possible to keep the original height of the molecules. (3) Furthermore, we compared differences of height and width between α2M and myosin filament. The result suggested that α2M had a disk‐like rather than a spherical form. Large proteins such as α2M are still difficult to crystallize for x‐ray analysis, and we tested the AFM method for the study of minute height differences of such proteins.