Abstract
CXCR1, a member in G-protein coupled receptor (GPCR) family, binds to chemokine interleukin-8 (IL-8) specifically and transduces signals to mediate immune and inflammatory responses. Despite the importance of CXCR1, high-resolution structure determination is hindered by the challenges in crystallization. It has been shown that properly designed mutants with enhanced thermostability, together with fusion partner proteins, can be useful to form crystals for GPCR proteins. In this study, in silico protein design was carried out by using homology modeling and molecular dynamics simulations. To validate the computational modeling results, the thermostability of several mutants and the wild type were measured experimentally. Both computational results and experimental data suggest that the mutant L126W has a significant improvement in the thermostability. This study demonstrated that in silico design can guide protein engineering and potentially facilitate protein crystallography research.
Highlights
Chemokine molecules and their interactions with receptors are crucial to cellular immunity, cancer and inflammation regulations[1,2]
We present the results on the study of the CXCR1-T4 lysozyme (T4L) complex, a G-protein coupled receptor (GPCR) protein with the T4L fusion partner, using homology modeling, molecular dynamics simulation and mutagenesis experiment methods
By comparing the computational results with experimental data, we found that using CCR2-T4L crystal structure as the homology template can yield a plausible predicted structure
Summary
Chemokine molecules and their interactions with receptors are crucial to cellular immunity, cancer and inflammation regulations[1,2]. The crystal structures of CXCR4 and CCR2 are very similar, with an RMSD (root mean square deviation) of less than 3 Å for TM regions, while the CXCR1 structure resolved using solid state NMR method has an RMSD of about 5 Å compared to either CXCR4 or CCR2 structures. These three structures share the same topology with the main differences in the loop regions and helix packing in the TM cores. Compared to the wild type, two mutants lead to enhanced thermostability, especially the L126W mutant improved the melting temperature by 8.37 °C
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