Revealing the assembly of filamentous proteins with scanning transmission electron microscopy

Autoři: Cristina Martinez-Torres aff001;  Federica Burla aff001;  Celine Alkemade aff001;  Gijsje H. Koenderink aff001
Působiště autorů: Department of Living Matter, AMOLF, Amsterdam, the Netherlands aff001;  Department of Bionanoscience, Kavli Institute of Nanoscience Delft, Faculty of Applied Sciences, Delft University of Technology, Delft, The Netherlands aff002
Vyšlo v časopise: PLoS ONE 14(12)
Kategorie: Research Article
doi: 10.1371/journal.pone.0226277


Filamentous proteins are responsible for the superior mechanical strength of our cells and tissues. The remarkable mechanical properties of protein filaments are tied to their complex molecular packing structure. However, since these filaments have widths of several to tens of nanometers, it has remained challenging to quantitatively probe their molecular mass density and three-dimensional packing order. Scanning transmission electron microscopy (STEM) is a powerful tool to perform simultaneous mass and morphology measurements on filamentous proteins at high resolution, but its applicability has been greatly limited by the lack of automated image processing methods. Here, we demonstrate a semi-automated tracking algorithm that is capable of analyzing the molecular packing density of intra- and extracellular protein filaments over a broad mass range from STEM images. We prove the wide applicability of the technique by analyzing the mass densities of two cytoskeletal proteins (actin and microtubules) and of the main protein in the extracellular matrix, collagen. The high-throughput and spatial resolution of our approach allow us to quantify the internal packing of these filaments and their polymorphism by correlating mass and morphology information. Moreover, we are able to identify periodic mass variations in collagen fibrils that reveal details of their axially ordered longitudinal self-assembly. STEM-based mass mapping coupled with our tracking algorithm is therefore a powerful technique in the characterization of a wide range of biological and synthetic filaments.

Klíčová slova:

Atomic force microscopy – Collagens – Extracellular matrix proteins – Microtubules – Molecular structure – Packing density – Protein structure – Tobacco mosaic virus


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2019 Číslo 12