BIOLOGICAL AND STRUCTURAL-PROPERTIES OF MIP-1-ALPHA EXPRESSED IN YEAST

The murine macrophage inflammatory proteins-1 alpha (MIP-1α) and MIP-1β are distinct but closely related cytokines. Partially purified mixtures of the two proteins affect neutrophil function and cause local inflammation and fever. The particular properties of MIP-1α have not been well studied, although it has been identified as being identical to an inhibitor of haemopoietic stem cell growth. We have expressed MIP-1α in yeast cells and purified it to sequence homogeneity. Structural analysis of this biologically active material by circular dichroism and fluorescence spectroscopy confirms that MIP-1α has a very similar secondary and tertiary structure to platelet factor 4 and interleukin 8 with which it shares limited sequence homology. The in-vitro stem cell inhibitory properties have been confirmed using a range of murine progenitor cells including purified bone marrow progenitor cells (FACS-1), the FDCP-mix A4 cell line, and spleen colony forming unit (CFU-S) populations. Plateau levels of inhibition of stem cell growth were achieved using concentrations of 0.15 μg/ml MIP-1α. We have also demonstrated that MIP-1α is active in vivo: 5 μg of MIP-1α per mouse given as a bolus injection, protects stem cells from subsequent in-vitro killing by tritiated thymidine. MIP-1α was also shown to enhance the proliferation of more committed progenitor granulocyte macrophage-colony forming cells (GM-CFC) in response to granulocyte macrophage-colony stimulating factor (GM-CSF). © 1992.