Dual energy landscape: The functional state of the β‐barrel outer membrane protein G molds its unfolding energy landscape

Abstract

We applied dynamic single-molecule force spectroscopy to quantify the parameters (free energy of activation and distance of the transition state from the folded state) characterizing the energy barriers in the unfolding energy landscape of the outer membrane protein G (OmpG) from Escherichia coli. The pH-dependent functional switching of OmpG directs the protein along different regions on the unfolding energy landscape. The two functional states of OmpG take the same unfolding pathway during the sequential unfolding of β-hairpins I-IV. After the initial unfolding events, the unfolding pathways diverge. In the open state, the unfolding of β-hairpin V in one step precedes the unfolding of β-hairpin VI. In the closed state, β-hairpin V and β-strand S11 with a part of extracellular loop L6 unfold cooperatively, and subsequently β-strand S12 unfolds with the remaining loop L6. These two unfolding pathways in the open and closed states join again in the last unfolding step of β-hairpin VII. Also, the conformational change from the open to the closed state witnesses a rigidified extracellular gating loop L6. Thus, a change in the conformational state of OmpG not only bifurcates its unfolding pathways but also tunes its mechanical properties for optimum function.

Locating an extracellular K+-dependent interaction site that modulates betaine-binding of the Na+-coupled betaine symporter BetP

“…The free unfolding energies, ΔG u , ranging between 21 to 28 k B T (Table 1) were similar to those reported in literature (22,33,(36)(37)(38). Most pronounced changes were observed again for EH2 in which the free unfolding energy dropped in the presence of K þ by 3-5 k B T. In contrast, the free unfolding energies of all other structural regions showed much smaller changes, which suggest that they were largely independent of the presence of K þ and betaine.…”

Molecular Plasticity of the Human Voltage-Dependent Anion Channel Embedded Into a Membrane

“…This includes the distance x u from the folded state to the transition state towards unfolding, which characterizes the width of the energy valley that stabilizes the folded structure. Our analysis revealed that x u values in hVDAC1 were ~30–50% larger than those determined for α-helices of most transmembrane proteins (Janovjak et al, 2008), and comparison with results for the transmembrane β-barrel of OmpG suggested the same trend (Damaghi et al, 2010). In addition, we obtained quantitative information about the mechanical rigidity of the hVDAC1 channel: its mechanical rigidity is comparable to that of different transmembrane α-helices (Kawamura et al, 2013; Zocher et al, 2012), but is a factor 2–10 below that of the β-strands of the transmembrane β-barrel protein OmpG (Damaghi et al, 2010).…”

Cholesterol increases kinetic, energetic, and mechanical stability of the human β2-adrenergic receptor

“…Measuring the kinetic response of the individual structural elements by dynamic SMFS (DFS) allows quantification of their unfolding free energy, kinetic stability, conformational variability, and mechanical flexibility (33). Both SMFS and DFS have been applied to characterize these parameters of various membrane proteins and follow how they change on changing their functional state or physiological environment (34)(35)(36)(37)(38)(39).…”