Heart failure is a major cause of death and disability. Impairments in blood circulation that accompany heart failure can be traced, in part, to alterations in the activity of the sarcoplasmic reticulum Ca2+ pump that are induced by its interactions with phospholamban, a reversible inhibitor. If phospholamban becomes superinhibitory or chronically inhibitory, contractility is diminished, inducing dilated cardiomyopathy in mice and humans. In mice, phospholamban seems to encumber an otherwise healthy heart, but humans with a phospholamban-null genotype develop early-onset dilated cardiomyopathy.

Phospholamban: a crucial regulator of cardiac contractility

Allosteric regulation of protein function is a mechanism by which an event in one place of a protein structure causes an effect at another site, much like the behavior of a telecommunications network in which a collection of transmitters, receivers and transceivers communicate with each other across long distances. For example, ligand binding or an amino acid mutation at an allosteric site can alter enzymatic activity or binding affinity in a distal region such as the active site or a second binding site. The mechanism of this site-to-site communication is of great interest, especially since allosteric effects must be considered in drug design and protein engineering. In this review, conformational mobility as the common route between allosteric regulation and catalysis is discussed. We summarize recent experimental data and the resulting insights into allostery within proteins, and we discuss the nature of future studies and the new applications that may result from increased understanding of this regulatory mechanism.

Allosteric regulation and catalysis emerge via a common route.

Cooperativity in Noncovalent Interactions.

Allostery in Disease and in Drug Discovery

Hippo-Independent Activation of YAP by the GNAQ Uveal Melanoma Oncogene through a Trio-Regulated Rho GTPase Signaling Circuitry

https://www.cell.com/biophysj/pdf/S0006-3495(03)74681-5.pdf

NMR Solution Structure and Topological Orientation of Monomeric Phospholamban in Dodecylphosphocholine Micelles

Allosteric signaling in proteins requires long-range communication mediated by highly conserved residues, often triggered by ligand binding. In this article, we map the allosteric network in the catalytic subunit of protein kinase A using NMR spectroscopy. We show that positive allosteric cooperativity is generated by nucleotide and substrate binding during the transitions through the major conformational states: apo, intermediate, and closed. The allosteric network is disrupted by a single site mutation (Y204A), which also decouples the cooperativity of ligand binding. Because protein kinase A is the prototype for the entire kinome, these findings may serve as a paradigm for describing long-range coupling in other protein kinases.

https://www.pnas.org/content/pnas/105/2/506.full.pdf

Allosteric cooperativity in protein kinase A.

Sarcolipin (SLN) is a 31 amino acid integral membrane protein that regulates Ca-ATPase activity in skeletal muscle. Here, we report the three-dimensional structure and topology of synthetic SLN in lipid environments, as determined by solution and solid-state NMR spectroscopy. 2D solution NMR experiments were performed on SLN solubilized in sodium dodecyl sulfate (SDS) micelles. We found that SLN adopts a highly defined R-helical conformation from F9 through R27, with a backbone RMSD of 0.65 A and a side chain RMSD of 1.66 A. The N-terminus (M1 through L8) and the C-terminus (S28 through Y31) are mostly unstructured. The orientation of the SLN was determined using one-dimensional 15 N NMR solid-state spectroscopy. The protein was incorporated into phospholipid bilayers prepared from a mixture of 1,2-dioleoyl-sn-glycero-3-phosphocholine and 1,2-dioleoyl-sn-glycero-3- phosphoethanolamine. The 15 N chemical shift solid-state spectra from selectively labeled SLN samples indicate that SLN orients perpendicularly to the plane of the membrane bilayers. These results support the proposed mechanism of Ca-ATPase regulation of SLN via protein-protein intramembranous interactions between the highly conserved transmembrane domains of the Ca-ATPase and the conserved transmembrane domain of SLN.

Structure and Orientation of Sarcolipin in Lipid Environments

Solid-state NMR spectroscopy, in conjunction with rigid body molecular dynamics calculations, shows that monomeric phospholamban in lipid bilayers has two distinct helical domains, with an interhel...

Solid-state NMR and rigid body molecular dynamics to determine domain orientations of monomeric phospholamban.

Human islet amyloid polypeptide (hIAPP), or amylin, is a 37 amino acid hormone secreted by pancreatic β-cells. hIAPP constitutes ∼90% of the amyloid deposits found in type II diabetic patients. It has been shown that the central region of the peptide (hIAPP20–29) constitutes the nucleation site for the amyloidogenic process with F23 playing a key role in the formation of the β-pleated structures. In addition, it has been proposed that an important stage in the cytotoxicity of hIAPP is its interaction with the β-cell membranes. As a first step toward the characterization of the interaction of hIAPP with cell membranes, we determined conformational preferences of hIAPP20–29 in membrane-mimicking environments. We found that upon interacting with negatively charged micelles, the dominant conformation of hIAPP20–29 is a distorted type I β-turn centered on residues F23 and G24, with F23, A25, and I26 forming a small hydrophobic cluster that may facilitate the interaction of this peptide with the membrane bilayer. Moreover, we were able to elucidate the topological orientation of the peptide that is absorbed on the micelle surface, with the hydrophobic cluster oriented toward the hydrocarbon region of the micelles and both N- and C-termini exposed to the solvent. © 2003 Wiley Periodicals, Inc. Biopolymers 69: 29–41, 2003

Alessandro Mascioni

Papers

NMR Solution Structure and Topological Orientation of Monomeric Phospholamban in Dodecylphosphocholine Micelles

Allosteric cooperativity in protein kinase A.

Structure and Orientation of Sarcolipin in Lipid Environments

Solid-state NMR and rigid body molecular dynamics to determine domain orientations of monomeric phospholamban.

Conformational preferences of the amylin nucleation site in SDS micelles: an NMR study.