Chromatography of PBi soluble lysin

Chromatography of PBi soluble lysin

Column is Pall Acrosep Q HypeD (quaternary ammonium). Two ml of sample 12 (PBi soluble lysin) was applied. The column was eluted by step gradients of ammonium acetate (2ml each) of 0.05, 0.1, 0.2, 0.3, and 0.5M. Fractions 21-26 represent 0.3M ammonium acetate and 26-30 0.5M ammonium acetate.

20 microliter aliquots were added to 100 microliters (50microliters pH 6 MES, 50 microliters 4MU-beta-NAc-glucosamidine) and incubated 10 min at 50C.

Beta-NAc-glucosaminidase assay of column fractions (earlier fractions had no activity)

As above, but with 30 microliter aliquots of column fractions and 4MU-alpha-D-glucoside as substrate

Tubes were photgraphed through a 520nm long pass filter (16" exposure)

Starting material was sample 12 from here. Note that glycosidases elute much later (0.3-0.5M ammonium acetate) than WAL lysin

N-Ac-beta-glucosaminidase is secreted by Chlorella!