Investigation of CC and CXC chemokine quaternary state mutants

The chemokine family forms two different types of homodimer despite members sharing nearly identical folds. To study the formation of quaternary structure in this family, rational mutagenesis was employed on a representative member of each subfamily (MIP-1beta and IL-8). The variants were studied by analytical ultracentrifugation and NMR, and it was determined that formation of a folded monomer from a natural chemokine dimer is reasonably facile, while conversion between dimer types is not. Monomeric variants of MIP-1beta and IL-8 were randomly mutated and a lambda phage-based selection system was employed in a novel way to screen for dimerization. A total of 6,000,000 random mutants were screened, but no dimers were formed, suggesting again that the chemokine fold is robust and amenable to sequence variation, while the chemokine dimer is much more difficult to attain. This work represents a biophysical analysis of an array of chemokine quaternary state variants.