Biology subjects

Rye, H.

Publications and source records attributed to Rye, H..

3 recordsLinked to original sources

bioRxiv2025

Single particle dynamics of protein aggregation and disaggregation in the presence of the sHsp proteins IbpAB

The small heat shock proteins (sHsps) are a key class of molecular chaperones that can inhibit protein aggregation and enhance protein recovery from aggregates. However, the mechanisms sHsps employ to carry out these roles are not well understood, in part because the highly heterogeneous and dynamic particles they form with aggregating proteins are difficult to study with traditional biophysical tools. Here we have applied a novel single particle fluorescence technique known as Burst Analysis Spectroscopy (BAS) to the study of the E. coli sHsps IbpA and IbpB (IbpAB). We show that in the presence of IbpAB, two different model proteins converge toward similar, limited aggregate particle size distributions. Additionally, while IbpAB dramatically accelerates the disassembly of protein aggregates by the bacterial KJEB bi-chaperone disaggregase, this enhancement does not appear to be strongly influenced by aggregate particle size. Rather, it is the ability of IbpAB to alter aggregate structure during particle formation that appears to be essential for stimulated disassembly. These observations support a model of aggregate recognition by IbpAB that is not only highly adaptable but capable of shaping aggregate particles into a specialized range of physical properties that are necessary for efficient protein disaggregation.

biochemistry↗

bioRxiv2024

GTP hydrolysis triggers membrane remodeling by AMPH-1

Membrane-enclosed transport carriers return biological molecules from the recycling endosome to the plasma membrane using a mechanism that is not well understood. In C. elegans, the formation of carriers from the recycling endosome requires the amphiphysin protein, AMPH-1. Recently, we found that purified AMPH-1 is sufficient for tubulation and vesiculation of liposomes in a mechanism that is regulated by guanine nucleotides. Here we propose a model linking GTP binding and hydrolysis to the membrane binding and tubulation required for transport carrier formation. We find that GTP binding stabilizes interactions between AMPH-1 and the membrane through amphipathic, N-terminal alpha helices, which are found at the tips of the arc-shaped, homodimeric structure. By contrast, in the post-hydrolysis, GDP-bound state, these helices are repositioned to interact with the N-terminal helices of other homodimers, to form an oligomeric AMPH-1 lattice that tubulates the membrane, in preparation for carrier formation by membrane fission.

biochemistry↗

bioRxiv2022

Binding of ATP to GroEL: a Case for Variable-Temperature Native Mass Spectrometry Thermodynamic Studies

Understanding how large, oligomeric protein complexes respond to the binding of small ligands and other proteins is essential for describing the molecular basis of life. This in turn requires a complete characterization of the binding energetics and correlation of thermodynamic data with interacting structures, including effects of small molecules and solvent. However, the size of many protein oligomers, the myriad intermediate ligation states they can populate, and their often complex allosteric regulation typically restrict analysis by traditional methods to low resolution, ensemble averages. Here, we employ variabletemperature electrospray ionization native mass spectrometry to determine the thermodynamics for stepwise binding of up to ATP molecules to the 801 kDa GroEL complex, a tetradecamer chaperonin complex. Binding thermodynamics reveal strong enthalpy-entropy compensation (EEC) and high degrees of cooperativity are observed for formation of GroEL-ATP7 and GroEL-ATP14. These are evidenced by entropically favored ATP binding to the cis ring (formation of GroEL-ATP1-7), with variations in EEC for subsequent binding of ATP to the trans ring (GroEL-ATP8-14), as expected for negative inter-ring cooperativity. Entropy driven ATP binding to the GroEL tetradecamer is consistent with ligand induced conformational changes of the GroEL tetradecamer, though the magnitude of the entropy change suggests that reorganization of GroEL-hydrating water molecules and/or expulsion of water from the GroEL cavity may also play a key role. By determining the thermodynamic signatures for individual ligand binding reactions to the large, nearly MDa GroEL complex, we expand our fundamental view of chaperonin functional chemistry. Moreover, this work and related studies of protein-ligand interactions illustrate unparalleled capabilities of vT-ESI-nMS for thermodynamics studies of protein interactions with ligands, and other molecules, such as proteins and drugs.

biophysics↗