During Dictyostelium development, glycogen degradation is a crucial event that
provides glucose monomers that are utilized to synthesize the essential structural
components for cellular differentiation. The degradation is catalyzed by the product of the
glycogen phosphorylase-2 gene. Cloning and sequencing of the gp-2 gene revealed several repeated sequences in the promoter region that are putative regulatory sites. I present here
the purification of a DNA binding protein that binds to the 3' "C" box sequence in the gp-2
promoter using a DEAE Sephacel resin and specific "C" DNA affinity column
chromatography. With undifferentiated amoebae cell extract, a DNA binding protein
migrated at 0.40 Rf and with 17 hr differentiated cell extract, the protein migrated at 0.32
Rf. Both the 0.32 and 0.40 Rf proteins were purified to homogeneity and showed to
consist of three subunits of 18 kD, 35 kD and 62 kD (for 0.40 Rf) or 81 kD (for 0.32 Rf).
Amino acid sequence analysis showed identity between a region of the 62 and 81 kD subunits. I conclude that the difference in the shifted 0.40 Rf and 0.32 Rf bands in EMSA
is due to the 62 and 81 k.D subunits. A southwestern blot analysis of the 17 hr cell extract
demonstrated that the DNA binding activity resides in the 81 kD polypeptide. The effect of
pH and phosphatase inhibitors on a "conversion" between the 0.32 and 0.40 Rf bands was
examined. The results suggest that phosphorylation may be involved in the "conversion"
reaction. This study suggests that the purified protein may be a trans-acting factor that is
involved in gp-2 regulation.
next to an author's name indicates that all files or directories associated with their ETD are accessible from the Virginia Tech campus network only.
![[VT]](http://scholar.lib.vt.edu/images/ETD-db/restricted.gif)
