An Interdisciplinary Investigation of Conformational Changes in Quasi-Crystalline Protein Array R-bodies in Response to pH
type: CreativeWork
description: R-bodies are ribbon-like protein polymers that undergo a dramatic conformational change from a tightly coiled form at neutral pH to an extended helical spiral at acidic pH. R-bodies were found in bacterial endosymbionts of paramecia, where their forceful extension causes vacuolar membranes to rupture contributing to a type of inter-paramecium warfare (Pond et al., 1989). Previous work has shown that R-body extension is fast, reversible, extremely robust, and tunable by directed evolution (Polka & Silver, 2016). However, it remains unknown how micron-scale changes in the conformation of an R-body ribbon arise from pH-induced changes in its nanoscale subunits. Here we use an interdisciplinary approach combining DIC microscopy, cryo-electron microscopy, atomic force microscopy, and hydrogen deuterium exchange to study individual, purified R-bodies in vitro. We show that R-body extension and contraction are highly cooperative and hysteretic processes with changes in the magnitude and direction of ribbon curvature as well as changes in ribbon thickness. Viewed en face, the R-body ribbon is a two-dimensional quasi-crystalline lattice with very small unit-cell dimensions (11.5 x 14.3 Å) that do not change with pH. Viewed edge-on, the ribbon has a laminar structure with five layers at neutral pH, two of which become indistinct at acidic pH. We show that the C-termini of the main constituent helical proteins, Reb A and Reb B, undergo large pH-dependent changes in accessibility for hydrogen-deuterium exchange, implying a transition from disordered at neutral pH to ordered helices at acidic pH. We propose this disordered-to-helical transition in the C-termini of Reb A and Reb B alters the tension within the concave side of the ribbon, driving changes in the local curvature of the ribbon to cause the extension process. Our findings provide a basis for understanding the mechanism of R-body extension, which may guide efforts to engineer R-bodies for novel drug delivery applicatio…
abstract: R-bodies are ribbon-like protein polymers that undergo a dramatic conformational change from a tightly coiled form at neutral pH to an extended helical spiral at acidic pH. R-bodies were found in bacterial endosymbionts of paramecia, where their forceful extension causes vacuolar membranes to rupture contributing to a type of inter-paramecium warfare (Pond et al., 1989). Previous work has shown that R-body extension is fast, reversible, extremely robust, and tunable by directed evolution (Polka & Silver, 2016). However, it remains unknown how micron-scale changes in the conformation of an R-body ribbon arise from pH-induced changes in its nanoscale subunits. Here we use an interdisciplinary approach combining DIC microscopy, cryo-electron microscopy, atomic force microscopy, and hydrogen deuterium exchange to study individual, purified R-bodies in vitro. We show that R-body extension and contraction are highly cooperative and hysteretic processes with changes in the magnitude and direction of ribbon curvature as well as changes in ribbon thickness. Viewed en face, the R-body ribbon is a two-dimensional quasi-crystalline lattice with very small unit-cell dimensions (11.5 x 14.3 Å) that do not change with pH. Viewed edge-on, the ribbon has a laminar structure with five layers at neutral pH, two of which become indistinct at acidic pH. We show that the C-termini of the main constituent helical proteins, Reb A and Reb B, undergo large pH-dependent changes in accessibility for hydrogen-deuterium exchange, implying a transition from disordered at neutral pH to ordered helices at acidic pH. We propose this disordered-to-helical transition in the C-termini of Reb A and Reb B alters the tension within the concave side of the ribbon, driving changes in the local curvature of the ribbon to cause the extension process. Our findings provide a basis for understanding the mechanism of R-body extension, which may guide efforts to engineer R-bodies for novel drug delivery applicatio…
author: Cai, Guangyang
language: en_US
keywords: membrane disruption, pH responsive, protein arrays, reb, refractile bodies, Biochemistry, Biophysics
date published: 2023-08-14
date modified: 2026-07-24T05:58:16Z
provider: University of Washington
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: University of Washington

PropertyValue
property ID: Handle
value: http://hdl.handle.net/1773/50236