Coaction of Electrostatic and Hydrophobic Interactions: Dynamic Constraints on Disordered TrkA Juxtamembrane Domain

2019 ◽  
Author(s):  
Zichen Wang ◽  
Huaxun Fan ◽  
Xiao Hu ◽  
John Khamo ◽  
Jiajie Diao ◽  
...  
Keyword(s):  
Single Molecule ◽  
Protein Function ◽  
Hydrophobic Interactions ◽  
Transmembrane Helix ◽  
Point Mutations ◽  
Salt Bridge ◽  
Receptor Activation ◽  
Membrane Dynamics ◽  
Juxtamembrane Domain ◽  
Joint Work

<p>The receptor tyrosine kinase family transmits signals into cell via a single transmembrane helix and a flexible juxtamembrane domain (JMD). Membrane dynamics makes it challenging to study the structural mechanism of receptor activation experimentally. In this study, we employ all-atom molecular dynamics with Highly Mobile Membrane-Mimetic to capture membrane interactions with the JMD of tropomyosin receptor kinase A (TrkA). We find that PIP<sub>2 </sub>lipids engage in lasting binding to multiple basic residues and compete with salt bridge within the peptide. We discover three residues insertion into the membrane, and perturb it through computationally designed point mutations. Single-molecule experiments indicate the contribution from hydrophobic insertion is comparable to electrostatic binding, and in-cell experiments show that enhanced TrkA-JMD insertion promotes receptor ubiquitination. Our joint work points to a scenario where basic and hydrophobic residues on disordered domains interact with lipid headgroups and tails, respectively, to restrain flexibility and potentially modulate protein function.</p>

scholarly journals Coaction of Electrostatic and Hydrophobic Interactions: Dynamic Constraints on Disordered TrkA Juxtamembrane Domain

2019 ◽  
Author(s):  
Zichen Wang ◽  
Huaxun Fan ◽  
Xiao Hu ◽  
John Khamo ◽  
Jiajie Diao ◽  
...  
Keyword(s):  
Single Molecule ◽  
Protein Function ◽  
Hydrophobic Interactions ◽  
Transmembrane Helix ◽  
Point Mutations ◽  
Salt Bridge ◽  
Receptor Activation ◽  
Membrane Dynamics ◽  
Juxtamembrane Domain ◽  
Joint Work

<p>The receptor tyrosine kinase family transmits signals into cell via a single transmembrane helix and a flexible juxtamembrane domain (JMD). Membrane dynamics makes it challenging to study the structural mechanism of receptor activation experimentally. In this study, we employ all-atom molecular dynamics with Highly Mobile Membrane-Mimetic to capture membrane interactions with the JMD of tropomyosin receptor kinase A (TrkA). We find that PIP<sub>2 </sub>lipids engage in lasting binding to multiple basic residues and compete with salt bridge within the peptide. We discover three residues insertion into the membrane, and perturb it through computationally designed point mutations. Single-molecule experiments indicate the contribution from hydrophobic insertion is comparable to electrostatic binding, and in-cell experiments show that enhanced TrkA-JMD insertion promotes receptor ubiquitination. Our joint work points to a scenario where basic and hydrophobic residues on disordered domains interact with lipid headgroups and tails, respectively, to restrain flexibility and potentially modulate protein function.</p>

Coaction of Electrostatic and Hydrophobic Interactions: Dynamic Structural Constraints on Disordered TrkA Juxtamembrane Domain

2019 ◽  
Author(s):  
Zichen Wang ◽  
Huaxun Fan ◽  
Xiao Hu ◽  
John Khamo ◽  
Jiajie Diao ◽  
...  
Keyword(s):  
Hydrophobic Interactions ◽  
Transmembrane Helix ◽  
Membrane Dynamics ◽  
Signaling Proteins ◽  
Structural Constraints ◽  
Juxtamembrane Domain ◽  
Hydrophobic Membrane ◽  
Anionic Lipids ◽  
Lipid Diffusion

<br>Effective transmembrane signaling is vital to the life of the cell. However, plasma membrane dynamics pose numerous challenges for signaling proteins and attempts to study them. The receptor tyrosine kinase family transmits signals via a single transmembrane helix and a flexible juxtamembrane domain (JMD). Detailed<br>studies of JMD-membrane interactions are challenging due to naturally slow lipid diffusion. We employ a combination of all-atom molecular dynamics of highly mobile membrane mimetics with in vitro and in cell experiments to capture critical roles of electrostatic and hydrophobic membrane-JMD interactions of<br>tropomyosin receptor kinase A (TrkA). We show that a conserved three-residue hydrophobic patch enables JMD anchoring in the membrane, a single residue deletion can dramatically reverse it, and a single charged to hydrophobic residue mutation can enhance binding affinity. The highly charged anionic lipids are effective<br>at promoting binding and restraining JMD flexibility. Our work highlights versatility of molecular controls of these interactions.<br>

scholarly journals Decoding the Molecular Effects of Atovaquone Linked Resistant Mutations on Plasmodium falciparum Cytb-ISP Complex in the Phospholipid Bilayer Membrane

2021 ◽  
Vol 22 (4) ◽  
pp. 2138
Author(s):  
Lorna Chebon-Bore ◽  
Taremekedzwa Allan Sanyanga ◽  
Colleen Varaidzo Manyumwa ◽  
Afrah Khairallah ◽  
Özlem Tastan Bishop
Keyword(s):  
Plasmodium Falciparum ◽  
Active Site ◽  
Protein Function ◽  
Hydrophobic Interactions ◽  
Point Mutations ◽  
Md Simulations ◽  
Resistance Mechanisms ◽  
Phospholipid Bilayer ◽  
Active Site Residues ◽  
Presence And Absence

Atovaquone (ATQ) is a drug used to prevent and treat malaria that functions by targeting the Plasmodium falciparum cytochrome b (PfCytb) protein. PfCytb catalyzes the transmembrane electron transfer (ET) pathway which maintains the mitochondrial membrane potential. The ubiquinol substrate binding site of the protein has heme bL, heme bH and iron-sulphur [2FE-2S] cluster cofactors that act as redox centers to aid in ET. Recent studies investigating ATQ resistance mechanisms have shown that point mutations of PfCytb confer resistance. Thus, understanding the resistance mechanisms at the molecular level via computational approaches incorporating phospholipid bilayer would help in the design of new efficacious drugs that are also capable of bypassing parasite resistance. With this knowledge gap, this article seeks to explore the effect of three drug resistant mutations Y268C, Y268N and Y268S on the PfCytb structure and function in the presence and absence of ATQ. To draw reliable conclusions, 350 ns all-atom membrane (POPC:POPE phospholipid bilayer) molecular dynamics (MD) simulations with derived metal parameters for the holo and ATQ-bound -proteins were performed. Thereafter, simulation outputs were analyzed using dynamic residue network (DRN) analysis. Across the triplicate MD runs, hydrophobic interactions, reported to be crucial in protein function were assessed. In both, the presence and absence of ATQ and a loss of key active site residue interactions were observed as a result of mutations. These active site residues included: Met 133, Trp136, Val140, Thr142, Ile258, Val259, Pro260 and Phe264. These changes to residue interactions are likely to destabilize the overall intra-protein residue communication network where the proteins’ function could be implicated. Protein dynamics of the ATQ-bound mutant complexes showed that they assumed a different pose to the wild-type, resulting in diminished residue interactions in the mutant proteins. In summary, this study presents insights on the possible effect of the mutations on ATQ drug activity causing resistance and describes accurate MD simulations in the presence of the lipid bilayer prior to conducting inhibitory drug discovery for the PfCytb-iron sulphur protein (Cytb-ISP) complex.

Identification of secretin, vasoactive intestinal peptide and glucagon binding sites: from chimaeric receptors to point mutations

2002 ◽  
Vol 30 (4) ◽  
pp. 437-441 ◽  
Author(s):  
M. Waelbroeck ◽  
J. Perret ◽  
P. Vertongen ◽  
M. Van Craenenbroeck ◽  
P. Robberecht
Keyword(s):  
Vasoactive Intestinal Peptide ◽  
Transmembrane Helix ◽  
Point Mutations ◽  
Receptor Activation ◽  
Side Chain ◽  
Extracellular Loop ◽  
Glucagon Receptor ◽  
In The Wild ◽  
Type Receptor ◽  
Receptor Conformation

We have identified two basic residues that are important for the recognition of secretin and vasoactive intestinal peptide (VIP) by their respective receptors. These two peptides containing an Asp residue at position 3 interacted with an arginine residue in transmembrane helix 2 (TM2) of the receptor, and the lysine residue in extracellular loop 1 (ECL1) stabilized the active receptor conformation induced by the ligand. The glucagon receptor possesses a Lys instead of an Arg in TM2, and an Ile instead of Lys in ECL1; it markedly prefers a Gln side chain in position 3 of the ligand. Our results suggested that, in the wild-type receptor, the Ile side chain prevented access to the TM2 Lys side chain, but oriented the glucagon Gln3 side chain to its proper binding site. In the double mutant, the ECL1 Lys allowed an interaction between negatively charged residues in position 3 of glucagon and the TM2 Arg, resulting in efficient receptor activation by [Asp3]glucagon as well as by glucagon.

scholarly journals Glutamine synthetase as a central element in hepatic glutamine and ammonia metabolism: novel aspects

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Benedikt Frieg ◽  
Boris Görg ◽  
Holger Gohlke ◽  
Dieter Häussinger
Keyword(s):  
Glutamine Synthetase ◽  
Protein Function ◽  
Nitrosative Stress ◽  
Point Mutations ◽  
Central Element ◽  
Ph Sensitive ◽  
Tyrosine Nitration ◽  
Ammonia Metabolism ◽  
Functional Consequences

Abstract Glutamine synthetase (GS) in the liver is expressed in a small perivenous, highly specialized hepatocyte population and is essential for the maintenance of low, non-toxic ammonia levels in the organism. However, GS activity can be impaired by tyrosine nitration of the enzyme in response to oxidative/nitrosative stress in a pH-sensitive way. The underlying molecular mechanism as investigated by combined molecular simulations and in vitro experiments indicates that tyrosine nitration can lead to a fully reversible and pH-sensitive regulation of protein function. This approach was also used to understand the functional consequences of several recently described point mutations of human GS with clinical relevance and to suggest an approach to restore impaired GS activity.

scholarly journals Screening marine algae metabolites as high-affinity inhibitors of SARS-CoV-2 main protease (3CLpro): an in silico analysis to identify novel drug candidates to combat COVID-19 pandemic

2020 ◽  
Vol 63 (1) ◽  
Author(s):  
Ghazala Muteeb ◽  
Adil Alshoaibi ◽  
Mohammad Aatif ◽  
Md. Tabish Rehman ◽  
M. Zuhaib Qayyum
Keyword(s):  
Hydrophobic Interactions ◽  
In Silico Analysis ◽  
Salt Bridge ◽  
Competitive Inhibitor ◽  
Binding Energies ◽  
Dynamics Simulation ◽  
In Vivo Studies ◽  
Energy Calculation

AbstractThe recent dissemination of SARS-CoV-2 from Wuhan city to all over the world has created a pandemic. COVID-19 has cost many human lives and created an enormous economic burden. Although many drugs/vaccines are in different stages of clinical trials, still none is clinically available. We have screened a marine seaweed database (1110 compounds) against 3CLpro of SARS-CoV-2 using computational approaches. High throughput virtual screening was performed on compounds, and 86 of them with docking score <  − 5.000 kcal mol−1 were subjected to standard-precision docking. Based on binding energies (< − 6.000 kcal mol−1), 9 compounds were further shortlisted and subjected to extra-precision docking. Free energy calculation by Prime-MM/GBSA suggested RC002, GA004, and GA006 as the most potent inhibitors of 3CLpro. An analysis of ADMET (Absorption, Distribution, Metabolism, Excretion, and Toxicity) properties of RC002, GA004, and GA006 indicated that only RC002 (callophysin A, from red alga Callophycus oppositifolius) passed Lipinski’s, Veber’s, PAINS and Brenk’s filters and displayed drug-like and lead-like properties. Analysis of 3CLpro-callophysin A complex revealed the involvement of salt bridge, hydrogen bonds, and hydrophobic interactions. callophysin A interacted with the catalytic residues (His41 and Cys145) of 3CLpro; hence it may act as a mechanism-based competitive inhibitor. Docking energy and docking affinity of callophysin A towards 3CLpro was − 8.776 kcal mol−1 and 2.73 × 106 M−1, respectively. Molecular dynamics simulation confirmed the stability of the 3CLpro-callophysin A complex. The findings of this study may serve as the basis for further validation by in vitro and in vivo studies.

scholarly journals Residues within the Transmembrane Domain of the Glucagon-Like Peptide-1 Receptor Involved in Ligand Binding and Receptor Activation: Modelling the Ligand-Bound Receptor

2011 ◽  
Vol 25 (10) ◽  
pp. 1804-1818 ◽  
Author(s):  
K. Coopman ◽  
R. Wallis ◽  
G. Robb ◽  
A. J. H. Brown ◽  
G. F. Wilkinson ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Structural Model ◽  
Transmembrane Domain ◽  
Transmembrane Helix ◽  
Receptor Activation ◽  
Agonist Affinity ◽  
Camp Generation ◽  
N Terminus ◽  
Glucagon Like Peptide ◽  
Glucagon Like Peptide 1

The C-terminal regions of glucagon-like peptide-1 (GLP-1) bind to the N terminus of the GLP-1 receptor (GLP-1R), facilitating interaction of the ligand N terminus with the receptor transmembrane domain. In contrast, the agonist exendin-4 relies less on the transmembrane domain, and truncated antagonist analogs (e.g. exendin 9–39) may interact solely with the receptor N terminus. Here we used mutagenesis to explore the role of residues highly conserved in the predicted transmembrane helices of mammalian GLP-1Rs and conserved in family B G protein coupled receptors in ligand binding and GLP-1R activation. By iteration using information from the mutagenesis, along with the available crystal structure of the receptor N terminus and a model of the active opsin transmembrane domain, we developed a structural receptor model with GLP-1 bound and used this to better understand consequences of mutations. Mutation at Y152 [transmembrane helix (TM) 1], R190 (TM2), Y235 (TM3), H363 (TM6), and E364 (TM6) produced similar reductions in affinity for GLP-1 and exendin 9–39. In contrast, other mutations either preferentially [K197 (TM2), Q234 (TM3), and W284 (extracellular loop 2)] or solely [D198 (TM2) and R310 (TM5)] reduced GLP-1 affinity. Reduced agonist affinity was always associated with reduced potency. However, reductions in potency exceeded reductions in agonist affinity for K197A, W284A, and R310A, while H363A was uncoupled from cAMP generation, highlighting critical roles of these residues in translating binding to activation. Data show important roles in ligand binding and receptor activation of conserved residues within the transmembrane domain of the GLP-1R. The receptor structural model provides insight into the roles of these residues.

scholarly journals Point Mutations in Transmembrane Helices 2 and 3 of ExbB and TolQ Affect Their Activities in Escherichia coli K-12

2004 ◽  
Vol 186 (13) ◽  
pp. 4402-4406 ◽  
Author(s):  
Volkmar Braun ◽  
Christina Herrmann
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Transmembrane Helix ◽  
Point Mutations ◽  
Transmembrane Helices ◽  
The Third ◽  
Form Part ◽  
K 12

ABSTRACT Replacement of glutamate 176, the only charged amino acid in the third transmembrane helix of ExbB, with alanine (E176A) abolished ExbB activity in all determined ExbB-dependent functions of Escherichia coli. Combination of the mutations T148A in the second transmembrane helix and T181A in the third transmembrane helix, proposed to form part of a proton pathway through ExbB, also resulted in inactive ExbB. E176 and T148 are strictly conserved in ExbB and TolQ proteins, and T181 is almost strictly conserved in ExbB, TolQ, and MotA.

scholarly journals An Intramolecular Salt Bridge in Bacillus thuringiensis Cry4Ba Toxin Is Involved in the Stability of Helix α-3, Which Is Needed for Oligomerization and Insecticidal Activity

2017 ◽  
Vol 83 (20) ◽  
Author(s):  
Sabino Pacheco ◽  
Isabel Gómez ◽  
Jorge Sánchez ◽  
Blanca-Ines García-Gómez ◽  
Mario Soberón ◽  
...  
Keyword(s):  
Bacillus Thuringiensis ◽  
Double Point ◽  
Single Point ◽  
Three Dimensional ◽  
Point Mutations ◽  
Salt Bridge ◽  
Dimensional Structure ◽  
Cry Toxins ◽  
Content Type ◽  
Domain I

ABSTRACT Bacillus thuringiensis three-domain Cry toxins kill insects by forming pores in the apical membrane of larval midgut cells. Oligomerization of the toxin is an important step for pore formation. Domain I helix α-3 participates in toxin oligomerization. Here we identify an intramolecular salt bridge within helix α-3 of Cry4Ba (D111-K115) that is conserved in many members of the family of three-domain Cry toxins. Single point mutations such as D111K or K115D resulted in proteins severely affected in toxicity. These mutants were also altered in oligomerization, and the mutant K115D was more sensitive to protease digestion. The double point mutant with reversed charges, D111K-K115D, recovered both oligomerization and toxicity, suggesting that this salt bridge is highly important for conservation of the structure of helix α-3 and necessary to promote the correct oligomerization of the toxin. IMPORTANCE Domain I has been shown to be involved in oligomerization through helix α-3 in different Cry toxins, and mutations affecting oligomerization also elicit changes in toxicity. The three-dimensional structure of the Cry4Ba toxin reveals an intramolecular salt bridge in helix α-3 of domain I. Mutations that disrupt this salt bridge resulted in changes in Cry4Ba oligomerization and toxicity, while a double point reciprocal mutation that restored the salt bridge resulted in recovery of toxin oligomerization and toxicity. These data highlight the role of oligomer formation as a key step in Cry4Ba toxicity.

scholarly journals Activation-induced Deaminase (AID)-directed Hypermutation in the Immunoglobulin Sμ Region

2002 ◽  
Vol 195 (4) ◽  
pp. 529-534 ◽  
Author(s):  
Hitoshi Nagaoka ◽  
Masamichi Muramatsu ◽  
Namiko Yamamura ◽  
Kazuo Kinoshita ◽  
Tasuku Honjo
Keyword(s):  
B Lymphocytes ◽  
Single Molecule ◽  
Dna Cleavage ◽  
Somatic Hypermutation ◽  
Point Mutations ◽  
Genetic Alterations ◽  
Variable Regions ◽  
Class Switch ◽  
Large Deletions ◽  
Activation Induced Deaminase

Somatic hypermutation (SHM) and class switch recombination (CSR) cause distinct genetic alterations at different regions of immunoglobulin genes in B lymphocytes: point mutations in variable regions and large deletions in S regions, respectively. Yet both depend on activation-induced deaminase (AID), the function of which in the two reactions has been an enigma. Here we report that B cell stimulation which induces CSR but not SHM, leads to AID-dependent accumulation of SHM-like point mutations in the switch μ region, uncoupled with CSR. These findings strongly suggest that AID itself or a single molecule generated by RNA editing function of AID may mediate a common step of SHM and CSR, which is likely to be involved in DNA cleavage.

Sign in / Sign up

Export Citation Format

Share Document