Novel use of an osmolyte to dissect multiple thermodynamic linkages in a chemokine ligand-receptor system.
We have used trimethylamine N-oxide (TMAO), a protecting osmolyte, to dissect the complex thermodynamic linkages involved in the interaction between the chemokine interleukin-8 (IL-8) and the N-domain of its receptor CXCR1. Our results show that TMAO induces folding in the CXCR1 receptor N-domain and that the N-domain upon folding binds ligand with higher affinity. This represents, to our knowledge, the smallest domain that has been shown to be folded in osmolyte. Using the phase diagram method to analyze this thermodynamic relationship graphically, we also observe that TMAO favors ligand dimerization and that the dimeric ligand binds the receptor domain with lower affinity. We have thus been able to dissect coupling among three distinct processes, receptor domain folding, ligand dimerization, and ligand-receptor domain binding in this chemokine-receptor system. We also observe that the affinity of the related chemokine, melanoma growth stimulatory activity (MGSA), increases concurrent with N-domain folding similar to IL-8 but shows more profound differences on ligand dimerization. These studies establish a novel and innovative use of osmolytes to dissect linkages among different processes and exploit the phase diagram as a tool to graphically represent and dissect complex thermodynamic relationships in biological systems. On the basis of our observations and earlier work, we discuss the relevance of ligand dimerization in chemokine regulation.
Rajagopalan
Lavanya
L
Department of Human Biological Chemistry and Genetics and Sealy Center for Structural Biology, The University of Texas Medical Branch, Galveston, Texas 77555-1055, USA.
Rösgen
Jörg
J
Bolen
David Wayne
DW
Rajarathnam
Krishna
K
D054360:Chemokine CXCL1; D019743:Chemokines, CXC / Q000737:chemistry; D019281:Dimerization; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000737:chemistry; D016209:Interleukin-8 / Q000737:chemistry*; D008744:Methylamines; D011485:Protein Binding; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D023062:Receptors, Interleukin-8A / Q000737:chemistry*; D013816:Thermodynamics
C475829:CXCL1 protein, human; D054360:Chemokine CXCL1; D019743:Chemokines, CXC; D036341:Intercellular Signaling Peptides and Proteins; D016209:Interleukin-8; D008744:Methylamines; D023062:Receptors, Interleukin-8A; C005855:trimethyloxamine