Conformational States and Thermodynamics of α-Lactalbumin Bound to Membranes: A Case Study of the Effects of pH, Calcium, Lipid Membrane Curvature and Charge

Chenal, Alexandre; Vernier, Grégory; Savarin, Philippe; Bushmarina, Natalia A.; Gèze, Annabelle; Guillain, Florent; Gillet, Daniel; Forge, Vincent

doi:10.1016/j.jmb.2005.04.036

Cited by 46 publications

(64 citation statements)

References 67 publications

(85 reference statements)

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“…The electrostatically modulated marginal thermodynamic barrier of BLA provides an explanation to its readiness to partially unfold into a MG-like state on interfacial contact with the negatively charged membrane surface. Furthermore, it explains the structural changes observed in membranebound BLA in response to differences in the overall charge, lipid mix, and/or fluidity of the host membrane (46,56,58). From the apparent reciprocity between biological function and special folding properties we conclude that BLA is an example of a marginal folding barrier selected by natural evolution.…”

Section: Discussionmentioning

confidence: 78%

Large-scale modulation of thermodynamic protein folding barriers linked to electrostatics

Halskau

Pérez-Jiménez

Ibarra‐Molero

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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natural selection ͉ homologous proteins ͉ structural pK shifts ͉ conformational ensembles ͉ differential scanning calorimetry C onventionally, the folding and function of single-domain proteins are described as two-state processes in analogy to elementary chemical reactions. The protein molecule is assumed to reside in either of two states: folded or active and unfolded or inactive, which interconvert by crossing a high free-energy barrier with transition-state-like kinetics. However, proteins can in principle exist in many different conformations or microstates because of their thousands of degrees of freedom. Such inherent complexity is best described by using low-dimensional freeenergy surfaces, which are obtained by projecting the solventaveraged energy as a function of atomic coordinates onto a few order parameters [the energy landscape approach (1)]. A freeenergy surface description includes the two-state folding model as a particular scenario, but is more general. In this approach barriers, basins of attraction, and even finer topographical details of the surface (i.e., roughness) emerge from detailed balancing between conformational entropy and the energy from stabilizing interactions (see refs. 2 and 3 for some specific examples). Therefore, surfaces that exhibit marginal barriers or are even completely barrierless appear as interesting alternatives to the two-state folding picture (1). These predictions have been confirmed experimentally in recent years (4-6).If the folding barriers are comparable to the thermal energy (RT), ensembles of conformations with an intermediate degree of structure become significantly populated and interconvert with diffusive dynamics. This realization opens a realm of possibilities for the experimental study of protein folding reactions and mechanisms (recently reviewed in ref. 7). Moreover, it also has important implications for protein function because the conformational fluctuations associated with marginal folding barriers could be exploited to modulate activity and/or synchronize the action of enzymes in sequential reactions (8). Examples of how this could be achieved have been recently explored for protein binding with the fly-cast model (9, 10) and related analyses (11), as well as for the optimization of allosteric coupling (12).Another important consequence of shallow free-energy surfaces is that their shape can be resolved in equilibrium experiments sensitive to the energy fluctuations associated with protein conformation, such as differential scanning calorimetry (DSC). Building on this idea, Muñoz and Sanchez-Ruiz have recently developed a phenomenological variable-barrier model for the analysis of DSC data (13). In this analysis, the DSC thermogram is fitted to an idealized (i.e., a Landau quartic polynomial) one-dimensional free-energy surface from which it is possible to estimate the barrier height and the population of conformational ensembles with an intermediate degree of structure (13). Although these folding barriers are intrinsically ''thermodynamic,'' ...

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Section: Discussionmentioning

confidence: 78%

Large-scale modulation of thermodynamic protein folding barriers linked to electrostatics

Halskau

Pérez-Jiménez

Ibarra‐Molero

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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“…The maximum emission wavelength of the intrinsic fluorescence spectrum (l max ), which is sensitive to the tryptophan (Trp) environment, has been used to monitor the pH-induced conformational changes and partitioning of aMb between buffer and LUVs. LUVs have been preferred to SUVs as models of membranes because they are more stable, and their curvature is closer to that of the membranes in cells (Lepore et al 1992;Epand 1998;Heerklotz and Epand 2001;Bigay et al 2003;Yoshida et al 2004;Chenal et al 2005). Figure 2 shows the pH dependence of l max for the protein in solution (triangles) and in the presence of anionic LUVs ( -LUVs) (circles).…”

Section: Resultsmentioning

confidence: 99%

“…They have shown that aMb interacts with lipid bilayers at neutral pH. The discrepancy probably arises from the use by these authors of SUV, sonicated lipid vesicles, which are characterized by a higher curvature, favoring hydrophobic effects within the solvent-membrane interface (Lepore et al 1992;Heerklotz and Epand 2001;Chenal et al 2005). This difference in behavior of aMb in the presence of lipid vesicles of various curvatures highlights that protein membrane interactions are governed by subtle electrostatic and hydrophobic effects.…”

Section: Discussionmentioning

confidence: 98%

Interactions of apomyoglobin with membranes: Mechanisms and effects on heme uptake

et al. 2007

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The last step of the folding reaction of myoglobin is the incorporation of a prosthetic group. In cells, myoglobin is soluble, while heme resides in the mitochondrial membrane. We report here an exhaustive study of the interactions of apomyoglobin with lipid vesicles. We show that apomyoglobin interacts with large unilamellar vesicles under acidic conditions, and that this requires the presence of negatively charged phospholipids. The pH dependence of apomyoglobin interactions with membranes is a two-step process, and involves a partially folded state stabilized at acidic pH. An evident role for the interaction of apomyoglobin with lipid bilayers would be to facilitate the uptake of heme from the outer mitochondrial membrane. However, heme binding to apomyoglobin is observed at neutral pH when the protein remains in solution, and slows down as the pH becomes more favorable to membrane interactions. The effective incorporation of soluble heme into apomyoglobin at neutral pH suggests that the interaction of apomyoglobin with membranes is not necessary for the heme uptake from the lipid bilayer. In vivo, however, the ability of apomyoglobin to interact with membrane may facilitate its localization in the vicinity of the mitochondrial membranes, and so may increase the yield of heme uptake. Moreover, the behavior of apomyoglobin in the presence of membranes shows striking similarities with that of other proteins with a globin fold. This suggests that the globin fold is well adapted for soluble proteins whose functions require interactions with membranes.

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“…According to the currently accepted "helical anchor" model, an amphipathic ␣-helix packs its aliphatic side against the hydrophobic protein core in the soluble state and rotates it to present this side to the membrane in the bound state while the charge side of the helix faces the lipid headgroup region (22)(23)(24)(25)(26). We have previously postulated that the amphipathic ␣2-helix is an important element to determine the binding of PimA to the membrane (11,14).…”

Section: Labeled Suv Typementioning

confidence: 99%

Molecular Basis of Membrane Association by the Phosphatidylinositol Mannosyltransferase PimA Enzyme from Mycobacteria

Rodrigo-Unzueta

Martínez

Comino

et al. 2016

Journal of Biological Chemistry

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Phosphatidyl-myo-inositol mannosyltransferase A (PimA) is an essential glycosyltransferase that initiates the biosynthetic pathway of phosphatidyl-myo-inositol mannoside, lipomannan, and lipoarabinomannan, which are key glycolipids/lipoglycans of the mycobacterial cell envelope. PimA belongs to a large family of membrane-associated glycosyltransferases for which the understanding of the molecular mechanism and conformational changes that govern substrate/membrane recognition and catalysis remains a major challenge. Here, we determined that PimA preferentially binds to negatively charged phosphatidylmyo-inositol substrate and non-substrate membrane model systems (small unilamellar vesicle) through its N-terminal domain, inducing an important structural reorganization of anionic phospholipids. By using a combination of single-point mutagenesis, circular dichroism, and a variety of fluorescence spectroscopy techniques, we determined that this interaction is mainly mediated by an amphipathic ␣-helix (␣2), which undergoes a substantial conformational change and localizes in the vicinity of the negatively charged lipid headgroups and the very first carbon atoms of the acyl chains, at the PimA-phospholipid interface. Interestingly, a flexible region within the N-terminal domain, which undergoes ␤-strand-to-␣-helix and ␣-helix-to-␤-strand transitions during catalysis, interacts with anionic phospholipids; however, the effect is markedly less pronounced to that observed for the amphipathic ␣2, likely reflecting structural plasticity/variability. Altogether, we propose a model in which conformational transitions observed in PimA might reflect a molten globule state that confers to PimA, a higher affinity toward the dynamic and highly fluctuating lipid bilayer.

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Conformational States and Thermodynamics of α-Lactalbumin Bound to Membranes: A Case Study of the Effects of pH, Calcium, Lipid Membrane Curvature and Charge

Cited by 46 publications

References 67 publications

Large-scale modulation of thermodynamic protein folding barriers linked to electrostatics

Large-scale modulation of thermodynamic protein folding barriers linked to electrostatics

Interactions of apomyoglobin with membranes: Mechanisms and effects on heme uptake

Molecular Basis of Membrane Association by the Phosphatidylinositol Mannosyltransferase PimA Enzyme from Mycobacteria

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