Angiotensin II cyclic analogs as tools to investigate ATR biased signaling mechanisms.

G protein coupled receptors (GPCRs) produce pleiotropic effects by their capacity to engage numerous signaling pathways once activated. Functional selectivity (also called biased signaling), where specific compounds can bring GPCRs to adopt conformations that enable selective receptor coupling to distinct signaling pathways, continues to be significantly investigated. However, an important but often overlooked aspect of functional selectivity is the capability of ligands such as angiotensin II (AngII) to adopt specific conformations that may preferentially bind to selective GPCRs structures.

Design, synthesis, and biological evaluation of CXCR4 ligands.

A combination of the CXCR4 inverse agonist T140 with N-terminal CXCL12 oligopeptides has produced the first nanomolar synthetic CXCR4 agonists. In these agonists, the inverse agonistic portion provides affinity whereas the N-terminal CXCL12 sequence induces receptor activation. Several CXCR4 crystal structures exist with either CVX15, an inverse agonist closely related to T140 and IT1t, a small molecule; we therefore attempted to produce another CXCL12 oligopeptide combination with IT1t.

Structure-Activity Relationship and Signaling of New Chimeric CXCR4 Agonists.

The CXCR4 receptor binds with meaningful affinities only CXCL12 and synthetic antagonists/inverse agonists. We recently described high affinity synthetic agonists for this chemokine receptor, obtained by grafting the CXCL12 N-terminus onto the inverse agonist T140. While those chimeric molecules behave as agonists for CXCR4, their binding and activation mode are unknown.

STARD6 on steroids: solution structure, multiple timescale backbone dynamics and ligand binding mechanism.

START domain proteins are conserved α/β helix-grip fold that play a role in the non-vesicular and intracellular transport of lipids and sterols. The mechanism and conformational changes permitting the entry of the ligand into their buried binding sites is not well understood. Moreover, their functions and the identification of cognate ligands is still an active area of research.

Identification of Distinct Conformations of the Angiotensin-II Type 1 Receptor Associated with the Gq/11 Protein Pathway and the β-Arrestin Pathway Using Molecular Dynamics Simulations.

Biased signaling represents the ability of G protein-coupled receptors to engage distinct pathways with various efficacies depending on the ligand used or on mutations in the receptor. The angiotensin-II type 1 (AT1) receptor, a prototypical class A G protein-coupled receptor, can activate various effectors upon stimulation with the endogenous ligand angiotensin-II (AngII), including the Gq/11 protein and β-arrestins. It is believed that the activation of those two pathways can be associated with distinct conformations of the AT1 receptor.

Mode of binding of the cyclic agonist peptide TC14012 to CXCR7: identification of receptor and compound determinants.

The chemokine receptor CXCR7 is an atypical CXCL12 receptor that, as opposed to the classical CXCL12 receptor CXCR4, signals preferentially via the β-arrestin pathway and does not mediate chemotaxis. We previously reported that the cyclic peptide TC14012, a potent CXCR4 antagonist, also engaged CXCR7, albeit with lower potency. Surprisingly, the compound activated the CXCR7-arrestin pathway.

Identification of transmembrane domain 1 & 2 residues that contribute to the formation of the ligand-binding pocket of the urotensin-II receptor.

The vasoactive urotensin-II (UII), a cyclic undecapeptide widely distributed in cardiovascular, renal and endocrine systems, specifically binds the UII receptor (UT receptor), a G protein-coupled receptor (GPCR). The involvement of this receptor in numerous pathophysiological conditions including atherosclerosis, heart failure, hypertension, renal impairment and diabetes potentially makes it an interesting therapeutic target. To elucidate how UII binds the UT receptor through the identification of specific residues in transmembrane domains (TM) one (TM1) and two (TM2) that are involved in the formation of the receptor's binding pocket, we used the substituted-cysteine accessibility method (SCAM).

Identification of transmembrane domain 3, 4 & 5 residues that contribute to the formation of the ligand-binding pocket of the urotensin-II receptor.

Urotensin-II (UII), a cyclic undecapeptide, selectively binds the urotensin-II receptor (UT receptor), a G protein-coupled receptor (GPCR) involved in cardiovascular effects and associated with numerous pathophysiological conditions including hypertension, atherosclerosis, heart failure, pulmonary hypertension and others. In order to identify specific residues in transmembrane domains (TM) three (TM3), four (TM4) and five (TM5) that are involved in the formation of the UT receptor binding pocket, we used the substituted-cysteine accessibility method (SCAM). Each residue in the F118((3.

Thermodynamic and solution state NMR characterization of the binding of secondary and conjugated bile acids to STARD5.

STARD5 is a member of the STARD4 sub-family of START domain containing proteins specialized in the non-vesicular transport of lipids and sterols. We recently reported that STARD5 binds primary bile acids. Herein, we report on the biophysical and structural characterization of the binding of secondary and conjugated bile acids by STARD5 at physiological concentrations.

Structure of the human angiotensin II type 1 (AT1) receptor bound to angiotensin II from multiple chemoselective photoprobe contacts reveals a unique peptide binding mode.

Breakthroughs in G protein-coupled receptor structure determination based on crystallography have been mainly obtained from receptors occupied in their transmembrane domain core by low molecular weight ligands, and we have only recently begun to elucidate how the extracellular surface of G protein-coupled receptors (GPCRs) allows for the binding of larger peptide molecules. In the present study, we used a unique chemoselective photoaffinity labeling strategy, the methionine proximity assay, to directly identify at physiological conditions a total of 38 discrete ligand/receptor contact residues that form the extracellular peptide-binding site of an activated GPCR, the angiotensin II type 1 receptor. This experimental data set was used in homology modeling to guide the positioning of the angiotensin II (AngII) peptide within several GPCR crystal structure templates.

Critical hydrogen bond formation for activation of the angiotensin II type 1 receptor.

G protein-coupled receptors contain selectively important residues that play central roles in the conformational changes that occur during receptor activation. Asparagine 111 (N111(3.35)) is such a residue within the angiotensin II type 1 (AT(1)) receptor.

Impact of guideline-consistent therapy on outcome of patients with healthcare-associated and community-acquired pneumonia.

Background: A new category of healthcare-associated pneumonia (HCAP) has been added in the most recent American Thoracic Society/Infectious Diseases Society of America (ATS/IDSA) guidelines, since multidrug-resistant (MDR) pathogens are more common in patients with HCAP than in those with community-acquired pneumonia (CAP). The optimal empirical management of patients with HCAP remains controversial and adherence to guidelines is inconsistent.

Methods: A retrospective cohort study of 3295 adults admitted for pneumonia in an academic centre of Canada, between 1997 and 2008.

Temperature dependent photolabeling of the human angiotensin II type 1 receptor reveals insights into its conformational landscape and its activation mechanism.

We present a photoaffinity labeling study of the human Angiotensin II (AngII) type 1 receptor (hAT(1)) and a constitutively active mutant (CAM) N111G hAT(1) at multiple temperatures using a p-benzoyl-l-phenylalanine (Bpa) containing AngII analogue (125)I-[Sar(1), Bpa(8)] AngII and the Methionine Proximity Approach (MPA). By introducing Met residues, which react selectively with Bpa, by mutagenesis in hAT(1) and its CAM, we were able to identify the position of residues that surround the Bpa moiety in the receptor-ligand complexes. Here we refined this characterization by controlling and varying (from -20 to 50 degrees C) the temperature at which the photolabeling was carried out.

A single-nucleotide polymorphism of alanine to threonine at position 163 of the human angiotensin II type 1 receptor impairs Losartan affinity.

Background And Objective: AT1 is the principal receptor for angiotensin II (AngII), which regulates blood pressure and osmotic homeostasis. Earlier studies have shown that position 163 interacts with the antihypertensive nonpeptide antagonist, Losartan. A recently discovered polymorphism found in humans (rs12721226) coding for residue 163 led us to determine whether this polymorphism would affect Losartan antihypertensive therapies.

Activation induces structural changes in the liganded angiotensin II type 1 receptor.

The octapeptide hormone angiotensin II (AngII) binds to and activates the human angiotensin II type 1 receptor (hAT(1)) of the G protein-coupled receptor class A family. Several activation mechanisms have been proposed for this family, but they have not yet been experimentally validated. We previously used the methionine proximity assay to show that 11 residues in transmembrane domain (TMD) III, VI, and VII of the hAT(1) receptor reside in close proximity to the C-terminal residue of AngII.

Photolabeling identifies transmembrane domain 4 of CXCR4 as a T140 binding site.

CXCR4, a G-protein-coupled receptor, which binds the chemokine stromal cell-derived factor 1 alpha (SDF-1alpha, CXCL12), is one of two co-receptors most frequently used by HIV-1 to infect CD4+ lymphocytes. The SDF-1alpha/CXCR4 axis is also involved in angiogenesis, in stem cell homing to bone marrow, in rheumatoid arthritis and in cancer. Here, we directly determined the binding site of the inverse agonist T140 on CXCR4 using photoaffinity labeling.