NMR structure and dynamics of monomeric neutrophil-activating peptide 2

Young, Helen C.; Roongta, Vikram; Daly, Thomas J.; Mayo, Kevin H.

doi:10.1042/bj3380591

Cited by 15 publications

(15 citation statements)

References 44 publications

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“…Further, NMR CSP-based methods have been shown to be extremely useful for describing residue-specific GAG interactions, as protein-GAG complexes are notoriously difficult to crystallize. The CXCL7 monomer chemical shift assignments were previously reported in the presence of 2-chloroethanol [16,57]. Our chemical shifts were similar, but not identical, and interestingly, our heteronuclear relaxation data of the monomer showed a structured 30s-loop, whereas previous studies carried out in the presence of chloroethanol showed substantial dynamics similar to those observed for terminal residues.…”

Section: Discussionsupporting

confidence: 83%

“…Comparison of our data to the previously reported relaxation data of CXCL7 in the presence of chloroethanol shows striking differences for the 30s-loop residues. Heteronuclear NOE measurements in the presence of 2-chloroethanol indicate a highly dynamic 30s-loop, especially residues Q33, V34 and E35, showing very low NOE values observed for the very terminal residues, whereas our values are similar to those of structured residues [16,31,32]. These observations suggest that chloroethanol influences dynamic properties, and so, these data may not fully reflect the dynamics of the native protein.…”

Section: Resultsmentioning

confidence: 62%

“…Our structure showed a backbone RMSD of 0.82 Å compared to the monomer units of the tetramer over the same regions, and differences in these regions are mainly due to extended β-strands and a slight change in the orientation of the helix. The largest differences between our monomer and the tetramer structure were observed for the N-terminus and 30s-loop residues, which can be attributed to their conformational flexibility [16,31,32] (Figure 3A). In general, we observed that the more dynamic a region, the greater the difference in its backbone RMSD.…”

Section: Resultsmentioning

confidence: 98%

“…A structure of the ethanol-induced monomer of CXCL7 has been previously reported, but its coordinates are not available in the protein data bank [16]. Generally, nuclear Overhauser effect (NOE)-driven structures require a sufficient number of long-range NOEs to describe different structural elements, their relative orientation and the global fold.…”

Section: Resultsmentioning

confidence: 99%

“…The structure of CXCL7 determined by crystallography corresponded to the tetrameric state [15], which is not surprising, as crystallography by its very nature results in the higher oligomeric state. The solution structure of a CXCL7 monomer determined in the presence of 2-chloroethanol that is known to disrupt intermolecular dimer and tetramer interactions has been reported, but its coordinates are not available in the public domain [16]. …”

Section: Introductionmentioning

confidence: 99%

See 4 more Smart Citations

Structural Basis of Native CXCL7 Monomer Binding to CXCR2 Receptor N-Domain and Glycosaminoglycan Heparin

Brown

Sepuru

Rajarathnam

2017

IJMS

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CXCL7, a chemokine highly expressed in platelets, orchestrates neutrophil recruitment during thrombosis and related pathophysiological processes by interacting with CXCR2 receptor and sulfated glycosaminoglycans (GAG). CXCL7 exists as monomers and dimers, and dimerization (~50 μM) and CXCR2 binding (~10 nM) constants indicate that CXCL7 is a potent agonist as a monomer. Currently, nothing is known regarding the structural basis by which receptor and GAG interactions mediate CXCL7 function. Using solution nuclear magnetic resonance (NMR) spectroscopy, we characterized the binding of CXCL7 monomer to the CXCR2 N-terminal domain (CXCR2Nd) that constitutes a critical docking site and to GAG heparin. We found that CXCR2Nd binds a hydrophobic groove and that ionic interactions also play a role in mediating binding. Heparin binds a set of contiguous basic residues indicating a prominent role for ionic interactions. Modeling studies reveal that the binding interface is dynamic and that GAG adopts different binding geometries. Most importantly, several residues involved in GAG binding are also involved in receptor interactions, suggesting that GAG-bound monomer cannot activate the receptor. Further, this is the first study that describes the structural basis of receptor and GAG interactions of a native monomer of the neutrophil-activating chemokine family.

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Section: Discussionsupporting

confidence: 83%

Section: Resultsmentioning

confidence: 62%

Section: Resultsmentioning

confidence: 98%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Structural Basis of Native CXCL7 Monomer Binding to CXCR2 Receptor N-Domain and Glycosaminoglycan Heparin

Brown

Sepuru

Rajarathnam

2017

IJMS

View full text Add to dashboard Cite

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Dynamics and thermodynamic properties of CXCL7 chemokine

et al. 2015

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Chemokines form a family of signaling proteins mainly responsible for directing the traffic of leukocytes, where their biological activity can be modulated by their oligomerization state. We characterize the dynamics and thermodynamic stability of monomer and homodimer structures of CXCL7, one of the most abundant platelet chemokines, using experimental methods that include circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy, and computational methods that include the anisotropic network model (ANM), molecular dynamics (MD) simulations and the distance constraint model (DCM). A consistent picture emerges for the effects of dimerization and Cys5-Cys31 and Cys7-Cys47 disulfide bonds formation. The presence of disulfide bonds is not critical for maintaining structural stability in the monomer or dimer, but the monomer is destabilized more than the dimer upon removal of disulfide bonds. Disulfide bonds play a key role in shaping the characteristics of native state dynamics. The combined analysis shows that upon dimerization flexibly correlated motions are induced between the 30s and 50s loop within each monomer and across the dimer interface. Interestingly, the greatest gain in flexibility upon dimerization occurs when both disulfide bonds are present, and the homodimer is least stable relative to its two monomers. These results suggest that the highly conserved disulfide bonds in chemokines facilitate a structural mechanism that is tuned to optimally distinguish functional characteristics between monomer and dimer.

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Structure–function studies of chemokine-derived carboxy-terminal antimicrobial peptides

Nguyen¹,

Chan²,

Boszhard³

et al. 2010

Biochimica et Biophysica Acta (BBA) - Biomembranes

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Recent reports which show that several chemokines can act as direct microbicidal agents have drawn renewed attention to these chemotactic signalling proteins. Here we present a structure-function analysis of peptides derived from the human chemokines macrophage inflammatory protein-3alpha (MIP-3alpha/CCL20), interleukin-8 (IL-8), neutrophil activating protein-2 (NAP-2) and thrombocidin-1 (TC-1). These peptides encompass the C-terminal alpha-helices of these chemokines, which have been suggested to be important for the direct antimicrobial activities. Far-UV CD spectroscopy showed that the peptides are unstructured in aqueous solution and that a membrane mimetic solvent is required to induce a helical secondary structure. A co-solvent mixture was used to determine solution structures of the peptides by two-dimensional (1)H-NMR spectroscopy. The highly cationic peptide, MIP-3alpha(51-70), had the most pronounced antimicrobial activity and displayed an amphipathic structure. A shorter version of this peptide, MIP-3alpha(59-70), remained antimicrobial but its structure and mechanism of action were unlike that of the former peptide. The NAP-2 and TC-1 proteins differ in their sequences only by the deletion of two C-terminal residues in TC-1, but intact TC-1 is a very potent antimicrobial while NAP-2 is inactive. The corresponding C-terminal peptides, NAP-2(50-70) and TC-1(50-68), had very limited and no bactericidal activity, respectively. This suggests that other regions of TC-1 contribute to its bactericidal activity. Altogether, this work provides a rational structural basis for the biological activities of these peptides and proteins and highlights the importance of experimental characterization of peptide fragments as distinct entities because their activities and structural properties may differ substantially from their parent proteins.

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NMR structure and dynamics of monomeric neutrophil-activating peptide 2

Cited by 15 publications

References 44 publications

Structural Basis of Native CXCL7 Monomer Binding to CXCR2 Receptor N-Domain and Glycosaminoglycan Heparin

Structural Basis of Native CXCL7 Monomer Binding to CXCR2 Receptor N-Domain and Glycosaminoglycan Heparin

Dynamics and thermodynamic properties of CXCL7 chemokine

Structure–function studies of chemokine-derived carboxy-terminal antimicrobial peptides

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