[52] Mannosidostreptomycin hydrolase

This chapter discusses the assay procedure and properties of mannosidostreptomycin hydrolase. Mannosidostreptomycin hydrolase is an enzyme that transforms mannosidostreptomycin to streptomycin by the hydrolytic removal of the mannose moiety. The two antibiotics are made concurrently during the course of fermentation by strains of Streptomyces griseus. The hydrolase is an inducible enzyme whose formation is subject to catabolite repression so that it is usually synthesized late in fermentation when the repressible carbon source is near depletion. The α-D-mannosidase enzyme is most conveniently assayed by use of the chromogenic substrate p-nitrophenyl-α-D-mannopyranoside. The p-nitrophenol liberated is determined spectrophotometrically at 400 nm. Streptomyces mannosidase hydrolyzes phenyl-α-D-mannopyranoside, mannosidostreptomycin, mannosidodihydrostreptomyein in addition to p-nitrophenyl-α-D-mannopyranoside. The Streptomyces enzyme is bound to the cell until lysis occurs. The cell-bound enzyme can be extracted into water, but its release is inhibited by sodium chloride, phosphate, or Tris.