Figure 4. Characterization of HS from HEK293 cells expressing mutant Hsepi.

(A) Western blot analysis of cell lysates (40 μg) shows overexpression of the mutant (Y168A point mutation) Hsepi, at a level comparable to that of Clone 9. (B) Epimerase activity of the mutant clone, assessed by ³H release upon incubation of cell lysates with labeled substrate, does not differ from that of the Mock clone. (C) Q-RT-PCR analysis confirms overexpression of Glce mRNA in the mutant clone. (D) Gel chromatography of ³⁵S-labeled HS on a Superose-6 column shows no difference in chain length of HS isolated from Y168A and Mock clones.