Mass Mapping Of Macromolecular Assemblies

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Scanning transmission electron microscopy (STEM) provides a versatile method for determining the molecular mass and hence the arrangement of subunits in large protein assemblies. Macromolecules are adsorbed onto a thin support film and a nanometer-sized electron probe is scanned across the specimen while the elastic-scattering signal is collected. The resulting digital image intensity is proportional to the local mass density of the specimen. Images can be recorded at low electron dose without significant radiation damage to the structures of interest. We have used this approach to measure mass per area of membranes extracted from the simple bacterium, Spiroplasma, as well as its contractile cytoskeleton, with the aim of constructing an inventory of the cellular components, which is of interest in the study of motility. We have tested the feasibility of extending the mass mapping approach in our 100 kV field-emission STEM by imaging undecagold clusters, each containing eleven gold atoms, adsorbed onto a thin carbon support. We are now investigating the application of monomaleimide undecagold to label specific proteins containing free sulfhydryl groups embedded in membranes that are labeled by undecagold attached to lipids.