scholarly journals Cytochrome c: A Multifunctional Protein Combining Conformational Rigidity with Flexibility

2014 ◽  
Vol 2014 ◽  
pp. 1-28 ◽  
Author(s):  
Reinhard Schweitzer-Stenner
Keyword(s):  
Protein Interactions ◽  
Redox Reactions ◽  
Biological Functions ◽  
Heme Group ◽  
Multifunctional Protein ◽  
Out Of Plane ◽  
Partial Unfolding ◽  
Partially Unfolded ◽  
Partially Unfolded States ◽  
Folding State

Cytochrome has served as a model system for studying redox reactions, protein folding, and more recently peroxidase activity induced by partial unfolding on membranes. This review illuminates some important aspects of the research on this biomolecule. The first part summarizes the results of structural analyses of its active site. Owing to heme-protein interactions the heme group is subject to both in-plane and out-of-plane deformations. The unfolding of the protein as discussed in detail in the second part of this review can be induced by changes of pH and temperature and most prominently by the addition of denaturing agents. Both the kinetic and thermodynamic folding and unfolding involve intermediate states with regard to all unfolding conditions. If allowed to sit at alkaline pH (11.5) for a week, the protein does not return to its folding state when the solvent is switched back to neutral pH. It rather adopts a misfolded state that is prone to aggregation via domain swapping. On the surface of cardiolipin containing liposomes, the protein can adopt a variety of partially unfolded states. Apparently, ferricytochrome c can perform biological functions even if it is only partially folded.

scholarly journals E3 Ubiquitin Ligase SPL2 Is a Lanthanide-Binding Protein

2021 ◽  
Vol 22 (11) ◽  
pp. 5712
Author(s):  
Michał Tracz ◽  
Ireneusz Górniak ◽  
Andrzej Szczepaniak ◽  
Wojciech Białek
Keyword(s):  
Ubiquitin Ligase ◽  
Conformational Changes ◽  
Protein Interactions ◽  
E3 Ubiquitin Ligase ◽  
Lanthanide Ions ◽  
E3 Ligases ◽  
Fluorescence Measurements ◽  
Partial Unfolding

The SPL2 protein is an E3 ubiquitin ligase of unknown function. It is one of only three types of E3 ligases found in the outer membrane of plant chloroplasts. In this study, we show that the cytosolic fragment of SPL2 binds lanthanide ions, as evidenced by fluorescence measurements and circular dichroism spectroscopy. We also report that SPL2 undergoes conformational changes upon binding of both Ca2+ and La3+, as evidenced by its partial unfolding. However, these structural rearrangements do not interfere with SPL2 enzymatic activity, as the protein retains its ability to auto-ubiquitinate in vitro. The possible applications of lanthanide-based probes to identify protein interactions in vivo are also discussed. Taken together, the results of this study reveal that the SPL2 protein contains a lanthanide-binding site, showing for the first time that at least some E3 ubiquitin ligases are also capable of binding lanthanide ions.

scholarly journals Mechanism of the small ATP-independent chaperone Spy is substrate specific

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Rishav Mitra ◽  
Varun V. Gadkari ◽  
Ben A. Meinen ◽  
Carlo P. M. van Mierlo ◽  
Brandon T. Ruotolo ◽  
...  
Keyword(s):  
Relative Affinity ◽  
Complex Protein ◽  
Partially Unfolded ◽  
Substrate Proteins ◽  
Partially Unfolded States

AbstractATP-independent chaperones are usually considered to be holdases that rapidly bind to non-native states of substrate proteins and prevent their aggregation. These chaperones are thought to release their substrate proteins prior to their folding. Spy is an ATP-independent chaperone that acts as an aggregation inhibiting holdase but does so by allowing its substrate proteins to fold while they remain continuously chaperone bound, thus acting as a foldase as well. The attributes that allow such dual chaperoning behavior are unclear. Here, we used the topologically complex protein apoflavodoxin to show that the outcome of Spy’s action is substrate specific and depends on its relative affinity for different folding states. Tighter binding of Spy to partially unfolded states of apoflavodoxin limits the possibility of folding while bound, converting Spy to a holdase chaperone. Our results highlight the central role of the substrate in determining the mechanism of chaperone action.

scholarly journals Chondroitin Sulfate/Dermatan Sulfate-Protein Interactions and Their Biological Functions in Human Diseases: Implications and Analytical Tools

2021 ◽  
Vol 9 ◽  
Author(s):  
Bin Zhang ◽  
Lianli Chi
Keyword(s):  
Cell Surface ◽  
Chondroitin Sulfate ◽  
Protein Interactions ◽  
Dermatan Sulfate ◽  
Interaction Analysis ◽  
Structural Complexity ◽  
Protein Characterization ◽  
Biological Functions ◽  
Gag Protein ◽  
Core Proteins

Chondroitin sulfate (CS) and dermatan sulfate (DS) are linear anionic polysaccharides that are widely present on the cell surface and in the cell matrix and connective tissue. CS and DS chains are usually attached to core proteins and are present in the form of proteoglycans (PGs). They not only are important structural substances but also bind to a variety of cytokines, growth factors, cell surface receptors, adhesion molecules, enzymes and fibrillary glycoproteins to execute series of important biological functions. CS and DS exhibit variable sulfation patterns and different sequence arrangements, and their molecular weights also vary within a large range, increasing the structural complexity and diversity of CS/DS. The structure-function relationship of CS/DS PGs directly and indirectly involves them in a variety of physiological and pathological processes. Accumulating evidence suggests that CS/DS serves as an important cofactor for many cell behaviors. Understanding the molecular basis of these interactions helps to elucidate the occurrence and development of various diseases and the development of new therapeutic approaches. The present article reviews the physiological and pathological processes in which CS and DS participate through their interactions with different proteins. Moreover, classic and emerging glycosaminoglycan (GAG)-protein interaction analysis tools and their applications in CS/DS-protein characterization are also discussed.

Abstract 434: A Novel 5-Step Model for the Mechanism of ABCA1-Mediated Nascent HDL Biogenesis

2012 ◽  
Vol 32 (suppl_1) ◽  
Author(s):  
Shuhui Wang ◽  
Gregory Brubaker ◽  
Kailash Gulshan ◽  
Jonathan D Smith
Keyword(s):  
Plasma Membrane ◽  
Membrane Lipids ◽  
Sodium Taurocholate ◽  
Hek293 Cells ◽  
Membrane Remodeling ◽  
Fluorescent Indicators ◽  
Step Model ◽  
Partial Unfolding ◽  
Cell Membrane Protein ◽  
Partially Unfolded

Objective— Lipid-poor apoA-I acts as an acceptor for cell cholesterol and phospholipids via the cell membrane protein ABCA1, generating nascent HDL. However, the mechanism of this process is not understood at the molecular level. Methods and Results— We propose a novel five-step model of nascent HDL biogenesis: ABCA1 remodeling of the plasma membrane lipids exposing phosphatidylserine and apoA-I binding to ABCA1 are the first two independent steps; third, ABCA1 facilitates apoA-I partial unfolding; forth, partially unfolded apoA-I inserts into the modified plasma membrane resulting in apoA-I lipidation; and fifth, nascent HDL is released from the cell. We created fluorescent apoA-I indicators that can monitor apoA-I unfolding and lipidation states. In cell free assays of reconstituted HDL (rHDL) generation from apoAI and DMPC liposomes, the fluorescent indicators demonstrated apoA-I unfolding and lipidation concurrent with rHDL formation. Next, HEK293 cells were stably transfected with different ABCA1 vectors encoding wild type (WT) and W590S and C1477R Tangier disease mutation isoforms. WT ABCA1 mediated cholesterol efflux to apoA-I (requires all steps) and sodium taurocholate (NaTC, requires only the membrane remodeling step,). Although neither mutant could efflux cholesterol efficiently to apoA-I, they were blocked at different steps. The W590S mutant bound apoAI but could not efflux cholesterol to NaTC, thus it was blocked at the membrane remodeling step. However, the C1477R mutant could not bind apoAI but could efflux cholesterol to NaTC, thus its activity was blocked at the apoAI binding step. When the lipidation indicator apoA-I was incubated with stably transfected HEK cells, we observed cell associated lipidated apoA-I in cells expressing WT ABCA1, but mostly unlipidated apoA-I was associated with the cells expressing W590S ABCA1. Conclusion— Our results support a novel five-step model for nascent HDL biogenesis: 1, 2) ABCA1 remodeling of the plasma membrane and apoA-I binding to ABCA1, which facilitate 3) apoA-I partial unfolding and 4) and lipidation by the remodeled membrane, followed by 5) the release of nascent HDL.

scholarly journals Relative Stability of the Scleroglucan Triple-Helix and Single Strand: an Insight from Computational and Experimental Techniques

2016 ◽  
Vol 230 (9) ◽  
Author(s):  
Gianfranco Bocchinfuso ◽  
Claudia Mazzuca ◽  
Paolo Conflitti ◽  
Davide Cori ◽  
Tommasina Coviello ◽  
...  
Keyword(s):  
Relative Stability ◽  
Triple Helix ◽  
Hydrophobic Interactions ◽  
Single Strand ◽  
Single Chain ◽  
Ph Values ◽  
Partial Unfolding ◽  
Triplex Structure ◽  
Partially Unfolded ◽  
The Stability

AbstractScleroglucan (Sclg) is a polysaccharide that exhibits a triple helix conformation (triplex), both in aqueous solution and in the solid state, which is lost in DMSO solution, at high temperature and at high pH values. The triplex conformation is characterized by a high rigidity, responsible of Sclg peculiar properties. Although the relative stability of triplex and single strand has already been investigated, different structural details are still missing.In the present study, we analyse the structural properties and the factors stabilizing the single chain and the triple helix of Sclg in different conditions. To this end, we simulated both systems in water and in DMSO. The triple helix has been also simulated in the presence of chemical damages on one of the three strands (to reproduce in silico the effect of sonication) or by inducing a partial unfolding of the triplex structure. The computational results have been compared with experimental evidences in which the triplex denaturation, at alkaline pH values, has been followed by monitoring the UV and CD spectra of Congo red, used as a probe molecule. Our results indicate that sonication breaks the Sclg chains without appreciably changing the stability of the other tracts of triple helix. The simulated perturbed or partially unfolded triplexes show a clear tendency to form less ordered aggregates. Finally, our simulations put in evidence an important role of the hydrophobic interactions both in the triplex stability and in the aggregation processes observed after induced denaturation.

scholarly journals Conserved salt-bridge competition triggered by phosphorylation regulates the protein interactome

2017 ◽  
Vol 114 (51) ◽  
pp. 13453-13458 ◽  
Author(s):  
John J. Skinner ◽  
Sheng Wang ◽  
Jiyoung Lee ◽  
Colin Ong ◽  
Ruth Sommese ◽  
...  
Keyword(s):  
G Protein ◽  
Protein Interactions ◽  
Salt Bridge ◽  
G Protein Coupled Receptor ◽  
Inhibitory Protein ◽  
Raf Kinase ◽  
Protein Interfaces ◽  
Partial Unfolding ◽  
Direct Recognition ◽  
G Protein Coupled

Phosphorylation is a major regulator of protein interactions; however, the mechanisms by which regulation occurs are not well understood. Here we identify a salt-bridge competition or “theft” mechanism that enables a phospho-triggered swap of protein partners by Raf Kinase Inhibitory Protein (RKIP). RKIP transitions from inhibiting Raf-1 to inhibiting G-protein–coupled receptor kinase 2 upon phosphorylation, thereby bridging MAP kinase and G-Protein–Coupled Receptor signaling. NMR and crystallography indicate that a phosphoserine, but not a phosphomimetic, competes for a lysine from a preexisting salt bridge, initiating a partial unfolding event and promoting new protein interactions. Structural elements underlying the theft occurred early in evolution and are found in 10% of homo-oligomers and 30% of hetero-oligomers including Bax, Troponin C, and Early Endosome Antigen 1. In contrast to a direct recognition of phosphorylated residues by binding partners, the salt-bridge theft mechanism represents a facile strategy for promoting or disrupting protein interactions using solvent-accessible residues, and it can provide additional specificity at protein interfaces through local unfolding or conformational change.

Exploration of partially unfolded states of human α-lactalbumin by molecular dynamics simulation11Edited by B. Honig

2001 ◽  
Vol 306 (2) ◽  
pp. 329-347 ◽  
Author(s):  
Emanuele Paci ◽  
Lorna J. Smith ◽  
Christopher M. Dobson ◽  
Martin Karplus
Keyword(s):  
Molecular Dynamics ◽  
Partially Unfolded ◽  
Partially Unfolded States
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