The colony-stimulating factors were originally described as biological activities in crude cell-conditioned media. Their intrinsic stability was a great advantage in their analysis and purification, but their heterogeneity, due primarily to differential glycosylation, and their extremely low concentration ensured that the purification of the CSFs would be extremely difficult and take up the better part of a decade. Even then, insufficient amounts of the CSFs were available from natural sources to allow detailed structural analysis and studies in vivo of their biological activities. With the advent of recombinant CSFs produced in Escherichia coli, it became apparent that glycosylation was not essential either for correct structure formation or for biological activity. This made it possible to perform detailed structural analyses of the CSFs as well as animal and clinical studies. The CSFs are all glycoproteins (Table 4.1), and despite having very little primary amino acid sequence homology, they have very similar folder structures; their stability and solubility are maintained by internal disulfide bonds (and intersubunit disulfide bonds in the case of M-CSF) and attached carbohydrate chains. In this chapter the purification, structural analysis, and structure-function studies of the CSFs will be described.

GM-CSF

The major CSF activity in medium conditioned by lung tissue from endotoxin-injected mice was recognized to have distinctive biological and biochemical properties (Sheridan and Metcalf, 1973a). This CSF was termed GM-CSF and was purified to apparent homogeneity by Burgess et al. (1977) using a combination of ammonium sulfate precipitation, ion-exchange chromatography, binding to concanavalin A-Sepharose, and gel filtration.