Elucidating the Molecular Interactions of Chemokine CCL2 Orthologs with Flavonoid Baicalin

Abstract

An integrated and controlled migration of leukocytes is necessary for the legitimate functioning and maintenance of the immune system. Chemokines and their receptors play a decisive role in regulating the leukocyte migration to the site of inflammation, a phenomena often referred to as chemotaxis. Chemokines and their receptors have become significant targets for therapeutic intervention considering their potential to regulate the immune system. Monocyte chemoattractant protein-1 (MCP-1/CCL2) is a preeminent member of CC chemokine family that facilitates crucial roles by orchestrating the recruitment of monocytes into inflamed tissues. Baicalin (BA), a major bioactive flavonoid, has been reported to attenuate chemokine-regulated leukocyte trafficking. However, no molecular details pertaining to its direct binding to chemokine(s)/receptor(s) are available till date. In the current study, using an array of monomers/dimers of human and murine CCL2 orthologs (hCCL2/mCCL2), we have shown that BA binds to the CCL2 protein specifically with nanomolar affinity ( K d = 270 ± 20 nM). NMR-based studies established that BA binds CCL2 in a specific pocket involving the N-terminal, β1- and β3-sheets. Docking studies suggested that the residues T16, N17, R18, I20, R24, K49, E50, I51, and C52 are majorly involved in complex formation through a combination of H-bonds and hydrophobic interactions. As the residues R18, R24, and K49 of hCCL2 are crucial determinants of monocyte trafficking through receptor/glycosaminoglycans (GAG) binding in CCL2 human/murine orthologs, we propose that baicalin engaging these residues in complex formation will result in attenuation of CCL2 binding to the receptor/GAGs, thus inhibiting the chemokine-regulated leukocyte trafficking.

Molecular Insights into Conformational Heterogeneity and Enhanced Structural Integrity of Helicobacter pylori DNA Binding Protein Hup at Low pH

“…Tyr fluorescence intensity maxima values at 306 nm were used to calculate the dissociation constants using the Stern−Volmer relationship and double log plots as described elsewhere. 48,49 Fluorescence Lifetime Spectroscopy. Fluorescence lifetime spectroscopy (FLS) measurements were performed on a time-correlated single-photon counting (TCSPC) spectrometer (Horiba Jobin Yvon Fluoro Cube Fluorescence Lifetime System) equipped with a Nano LED as pulse excitation source.…”

“…The ds-DNA thus formed was titrated at various concentrations to attain a final molar ratio of Hup with DNA in the range of 1:0.1 to 1:5. Tyr fluorescence intensity maxima values at 306 nm were used to calculate the dissociation constants using the Stern–Volmer relationship and double log plots as described elsewhere. , …”

“…As the diffusion properties are dependent on the molecular size and shape of a given protein, DOSY elucidates the presence of either dimer, monomer, and/or equilibrium of both. 49,65 In the present case, the DOSY results on the Hup protein at pH 6 and 2 were compared with those on standard proteins [SH3 domain7.2 kDa and hen egg lysozyme (HEL)14.3 kDa]. The diffusion profiles obtained at pH 2 were above HEL, thus representing for a molecular weight of ∼16−17 kDa (Figure S1).…”

Molecular Genetics and Cytotoxic Responses to Titanium Diboride and Zinc Borate Nanoparticles on Cultured Human Primary Alveolar Epithelial Cells

“…Moreover, in regards to providing modest antimutagenic potential, zinc borate was discovered to have moderate total antioxidant activity. With its antioxidative and antimutagenic qualities, zinc borate has the potential to be used in medicine [ 19 ].…”

Hydroxyl Groups on Annular Ring-B Dictate the Affinities of Flavonol–CCL2 Chemokine Binding Interactions

“… 47 , 48 On similar lines, baicalin (BA), a flavone-glycoside, was reported to modulate the chemotactic activity of CCL2 and other chemokines by interfering with the chemokine–receptor/GAG axis. 49 , 50 …”