EXPRESSION, PURIFICATION, AND CHARACTERIZATION OF A HIGHLY SOLUBLE N-TERMINAL-TRUNCATED FORM OF THE NEURON-SPECIFIC MEMBRANE-ASSOCIATED PHOSPHOPROTEIN SCG10

SCG10 is a neuron-specific growth-associated protein with high sequenc e homology to the ubiquitous phosphoprotein stathmin/Op18. The main st ructural difference between the two proteins is the 34-amino-acid N-te rminal extension of SCG10, which is responsible for the membrane attac hment, Full length SCG10 has been purified and shows limited solubilit y, in contrast to stathmin, which is a highly soluble protein, In orde r to obtain a more soluble form of SCG10 which would be better suited for biochemical and structural studies, we deleted the N-terminal exte nsion and expressed the C-terminal portion of the protein, Two forms o f N-terminal-truncated SCG10 (Delta SCG10 and Delta SCG10r) were purif ied to homogeneity in a four-step purification procedure, Delta SCG10 starts at amino acid 35 and Delta SCG10r at amino acid 48 in the SCG10 sequence, giving proteins of 16,899 and 15,189 kDa, respectively, The truncated SCG10 was highly soluble up to concentrations of 20 mg/ml, The proteins were like the full length SCG10 substrate for serine/thre onine protein kinases, including MAP kinase, PKA, and p34(cdc2) kinase , With these highly soluble forms of SCG10 biochemical and structural studies of this multiphosphoprotein become feasible. (C) 1997 Academic Press.