Dynamics-Derived Insights into Complex Formation between the CXCL8 Monomer and CXCR1 N-Terminal Domain: An NMR Study

TY - JOUR

T1 - Dynamics-Derived Insights into Complex Formation between the CXCL8 Monomer and CXCR1 N-Terminal Domain

T2 - An NMR Study

AU - Joseph, Prem Raj B.

AU - Spyracopoulos, Leo

AU - Rajarathnam, Krishna

PY - 2018/10/31

Y1 - 2018/10/31

N2 - Interleukin-8 (CXCL8), a potent neutrophil-activating chemokine, exerts its function by activating the CXCR1 receptor that belongs to class A G protein-coupled receptors (GPCRs). Receptor activation involves interactions between the CXCL8 N-terminal loop and CXCR1 N-terminal domain (N-domain) residues (Site-I) and between the CXCL8 N-terminal and CXCR1 extracellular/transmembrane residues (Site-II). CXCL8 exists in equilibrium between monomers and dimers, and it is known that the monomer binds CXCR1 with much higher affinity and that Site-I interactions are largely responsible for the differences in monomer vs. dimer affinity. Here, using backbone 15N-relaxation nuclear magnetic resonance (NMR) data, we characterized the dynamic properties of the CXCL8 monomer and the CXCR1 N-domain in the free and bound states. The main chain of CXCL8 appears largely rigid on the picosecond time scale as evident from high order parameters (S²). However, on average, S² are higher in the bound state. Interestingly, several residues show millisecond-microsecond (ms-μs) dynamics only in the bound state. The CXCR1 N-domain is unstructured in the free state but structured with significant dynamics in the bound state. Isothermal titration calorimetry (ITC) data indicate that both enthalpic and entropic factors contribute to affinity, suggesting that increased slow dynamics in the bound state contribute to affinity. In sum, our data indicate a critical and complex role for dynamics in driving CXCL8 monomer-CXCR1 Site-I interactions.

AB - Interleukin-8 (CXCL8), a potent neutrophil-activating chemokine, exerts its function by activating the CXCR1 receptor that belongs to class A G protein-coupled receptors (GPCRs). Receptor activation involves interactions between the CXCL8 N-terminal loop and CXCR1 N-terminal domain (N-domain) residues (Site-I) and between the CXCL8 N-terminal and CXCR1 extracellular/transmembrane residues (Site-II). CXCL8 exists in equilibrium between monomers and dimers, and it is known that the monomer binds CXCR1 with much higher affinity and that Site-I interactions are largely responsible for the differences in monomer vs. dimer affinity. Here, using backbone 15N-relaxation nuclear magnetic resonance (NMR) data, we characterized the dynamic properties of the CXCL8 monomer and the CXCR1 N-domain in the free and bound states. The main chain of CXCL8 appears largely rigid on the picosecond time scale as evident from high order parameters (S²). However, on average, S² are higher in the bound state. Interestingly, several residues show millisecond-microsecond (ms-μs) dynamics only in the bound state. The CXCR1 N-domain is unstructured in the free state but structured with significant dynamics in the bound state. Isothermal titration calorimetry (ITC) data indicate that both enthalpic and entropic factors contribute to affinity, suggesting that increased slow dynamics in the bound state contribute to affinity. In sum, our data indicate a critical and complex role for dynamics in driving CXCL8 monomer-CXCR1 Site-I interactions.

KW - 15N-relaxation

KW - chemokine

KW - CXCL8

KW - CXCR1

KW - dynamics

KW - G protein-coupled receptor (GPCR)

KW - isothermal titration calorimetry (ITC)

KW - nuclear magnetic resonance (NMR)

KW - receptor interactions

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U2 - 10.3390/molecules23112825

DO - 10.3390/molecules23112825

M3 - Article

C2 - 30384436

AN - SCOPUS:85055916876

VL - 23

JO - Molecules

JF - Molecules

SN - 1420-3049

IS - 11

ER -