Macromolecular crowding and intrinsically disordered proteins

Trosel, Yanitza (2023) Macromolecular crowding and intrinsically disordered proteins. Doctoral (PhD) thesis, Memorial University of Newfoundland.

[img] [English] PDF - Accepted Version
Available under License - The author retains copyright ownership and moral rights in this thesis. Neither the thesis nor substantial extracts from it may be printed or otherwise reproduced without the author's permission.

Download (46MB)


The intracellular milieu is crowded and heterogeneous, and this can have profound consequences for biomolecule motions and biochemical kinetics. Macromolecular crowding has been traditionally studied in artificial crowders like Ficoll and dextran or globular proteins such as bovine serum albumin. It is, however, not clear if the effects of artificial crowders on such phenomena are the same as the crowding that is experienced in a heterogeneous biological environment. Bacterial cells, for example, are composed of heterogeneous biomolecules with different sizes, shapes and charges. Using crowders composed of one of three different pretreatments of bacterial cell lysate (unmanipulated, ultracentrifuged, and anion exchanged), we examine the effects of crowding on the diffusivity of a model polymer and an intrinsically disordered protein (IDP). We measure the translational diffusivity, via diffusion NMR, of the test polymer polyethylene glycol (PEG) and the IDP α-synuclein in these bacterial cell lysates. We show that the small (Rg ~ 5 nm) test polymer shows a modest decrease in self-diffusivity with increasing crowder concentration for all lysate treatments. The corresponding self-diffusivity decrease in the artificial Ficoll crowder is much more pronounced. For the IDP, we show that α-synuclein has a significant decrease in the diffusivity in the presence of lysate crowders at low concentrations. Moreover, a comparison of the rheological response of biological and artificial crowders shows that while the artificial crowder Ficoll exhibits a Newtonian response even at high concentrations, the bacterial cell lysate is markedly non-Newtonian; it behaves like a shear-thinning uid with a yield stress. While at any concentration the rheological properties are sensitive to both lysate pretreatment and batch-to-batch variations, the PEG and α-synuclein diffusivity is nearly unaffected by the type of lysate pretreatment.

Item Type: Thesis (Doctoral (PhD))
Item ID: 16046
Additional Information: Includes bibliographical references (pages 95-108)
Keywords: macromolecular crowding, diffusion, rheology, bacterial cell lysate, diffusion NMR, polyethylene glycol, Ficoll
Department(s): Science, Faculty of > Physics and Physical Oceanography
Date: June 2023
Date Type: Submission
Digital Object Identifier (DOI):
Library of Congress Subject Heading: Macromolecules--Analysis; Rheology; Diffusion; Polyethylene glycol; Ficoll

Actions (login required)

View Item View Item


Downloads per month over the past year

View more statistics