Emerging Insights into the Structure and Function of Complement C5a Receptors

Pandey, Shubhi; Maharana, Jagannath; Li, Xaria X.; Woodruff, Trent M.; Shukla, Arun K.

doi:10.1016/j.tibs.2020.04.004

Cited by 63 publications

(80 citation statements)

References 80 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Three N-terminal tyrosine residues of C5aR2 may also be sulfated, but mutations of these residues did not significantly change C5a binding to C5aR2 34 , suggesting against the two-site binding mechanism for C5aR2. Our mechanistic insights into the C5a binding process to C5aR could offer a new framework to a better understanding or the intriguing functional divergence between the two C5a receptors and could potentially allow delineating the link between βarr conformational signatures and downstream functional outcomes 35 . Moreover, since a similar two-step two-site binding model has also been proposed for chemokine-receptor interactions 36,37 , our approach could also be useful to further study phenomena such as allosteric receptor interactions and ligand-biased receptor activation in this context as well.…”

Section: Discussionmentioning

confidence: 99%

“…For the 35 S-GTPγS binding assays, the membrane of HEK293S GnTI − cells expressing wtC5aR (~200 µg ml −1 ) or mutant variants was incubated with 200 nM purified G i protein for 30 min on ice in buffer containing 20 mM HEPES pH 7.5, 100 mM NaCl, 5 mM MgCl 2 , 3 μg ml −1 BSA, 0.1 μM TCEP, and 5 μM GDP to get the receptor and G i complex. Next, 25 μL aliquots of the pre-formed complex were mixed with 225 μL reaction buffer containing 20 mM HEPES, pH 7.5, 100 mM NaCl, 5 mM MgCl 2 , 3 μg ml −1 BSA, 0.1 μM TCEP, 1 μM GDP, 35 pM 35 S-GTPγS (Perkin Elmer), and C5a (R&D Systems).…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Submolecular probing of the complement C5a receptor–ligand binding reveals a cooperative two-site binding mechanism

et al. 2020

View full text Add to dashboard Cite

A current challenge to produce effective therapeutics is to accurately determine the location of the ligand-biding site and to characterize its properties. So far, the mechanisms underlying the functional activation of cell surface receptors by ligands with a complex binding mechanism remain poorly understood due to a lack of suitable nanoscopic methods to study them in their native environment. Here, we elucidated the ligand-binding mechanism of the human G protein-coupled C5a receptor (C5aR). We discovered for the first time a cooperativity between the two orthosteric binding sites. We found that the N-terminus C5aR serves as a kinetic trap, while the transmembrane domain acts as the functional site and both contributes to the overall high-affinity interaction. In particular, Asp282 plays a key role in ligand binding thermodynamics, as revealed by atomic force microscopy and steered molecular dynamics simulation. Our findings provide a new structural basis for the functional and mechanistic understanding of the GPCR family that binds large macromolecular ligands.

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Submolecular probing of the complement C5a receptor–ligand binding reveals a cooperative two-site binding mechanism

et al. 2020

View full text Add to dashboard Cite

show abstract

“…We next examined the potential activity of SB290157 on the other closely related complement receptor, C5aR2. As a non-canonical G-protein coupled receptor, C5aR2 does not couple to the common classes of G proteins and is devoid of the classical G protein-mediated signalling activities (26, 27). Human C5aR2 activation however recruits β-arrestins, which can be used as a readout of receptor activation (25, 28).…”

Section: Resultsmentioning

confidence: 99%

The ‘C3aR antagonist’ SB290157 is a partial C5aR2 agonist

Kumar

Lee

et al. 2020

Preprint

Self Cite

View full text Add to dashboard Cite

Innate immune complement activation generates the C3 and C5 protein cleavage products C3a and C5a, defined classically as anaphylatoxins. C3a activates C3a receptors (C3aR), while C5a activates two receptors (C5aR1 and C5aR2) to exert their immunomodulatory activities. The non-peptide compound, SB290157, was originally reported in 2001 as the first C3aR antagonist. In 2005, the first report on non-selective nature of SB290157 was published, where the compound exerted clear agonistic, not antagonistic, activity in variety of cells. Other studies also documented non-selective activities of this drug in vivo. These findings severely hamper data interpretation regarding C3aR when using this compound. Unfortunately, given the dearth of C3aR inhibitors, SB290157 still remains widely used to explore C3aR biology (>70 publications to date). Given these issues, in the present study we aimed to further explore SB290157's pharmacological selectivity by screening the drug against three human anaphylatoxin receptors, C3aR, C5aR1 and C5aR2, using transfected cells. We first confirmed that SB290157 acts as a potent agonist at human C3aR. We also identified that SB290157 exerts partial agonist activity at C5aR2 by mediating β-arrestin recruitment at higher compound doses. Notably, SB290157's activity at C5aR2 was more potent and efficacious than the current 'lead' C5aR2 agonist P32. Notwithstanding this, SB290157 showed inhibitory effect on C3a-mediated signalling in primary human macrophages. Our results therefore provide even more caution against using SB290157 as a research tool to explore C3aR function. Given the reported immunomodulatory and anti-inflammatory activities of C3aR and C5aR2 agonism, any function observed with SB290157 could be due to these off target activities.

show abstract

“…Dysfunction [ 19 ], neutrophil activation and neutrophil extracellular trap (NET) release [ 20 , 21 ], and complement system activation [ 22 ]. Emerging data suggest that C5a activation provides rapid protection from infectious challenge and can also transition to the ‘dark side’, becoming a driver or exacerbator of pathology in COVID-19-associated coagulopathy [ 10 , 23 ].…”

Section: Covid-19-associated Coagulopathy and Disease Severitymentioning

confidence: 99%

“…Although a detailed mechanism of C5aR2 signaling and its functional consequences remains to be elucidated, recent progress has shown that C5aR2 binds C5a and desarginated C5a (C5a-desArg) and internalizes them for intracellular degradation, thereby providing a negative regulatory mechanism to remove excess C5a from the circulation [ 25 ]. C5aR2 serves as a negative regulator and balancer of C5aR1 surface expression and helps to prevent overactivation of C5aR1 [ 23 ]. Additionally, C5aR2 induces robust high mobility group box 1 (HMGB1) production upon C5a stimulation associated with MEK1/2, JNK1/2, and PI3K/Akt activation [ 30 ].…”

Section: C5a and C5ars In Covid-19mentioning

confidence: 99%

The roles and potential therapeutic implications of C5a in the pathogenesis of COVID-19-associated coagulopathy

Liu

2021

Cytokine & Growth Factor Reviews

View full text Add to dashboard Cite

Emerging Insights into the Structure and Function of Complement C5a Receptors

Cited by 63 publications

References 80 publications

Submolecular probing of the complement C5a receptor–ligand binding reveals a cooperative two-site binding mechanism

Submolecular probing of the complement C5a receptor–ligand binding reveals a cooperative two-site binding mechanism

The ‘C3aR antagonist’ SB290157 is a partial C5aR2 agonist

The roles and potential therapeutic implications of C5a in the pathogenesis of COVID-19-associated coagulopathy

Contact Info

Product

Resources

About