CYTOCHEMICAL-LOCALIZATION OF N-ACETYL-BETA-D-GLUCOSAMINIDASE IN HYPHAE OF MUCOR-RACEMOSUS

Light microscopic localization of N-acetyl-β-d-glucosaminidase activity by the hydrolysis of α-naphthyl N-acetyl-β-d-glucosaminide and by the subsequent coupling of the released α-naphthol with Fast Red B and p-nitrobenzene diazonium tetrafluoroborate was obtained in Mucor racemosus. Enzyme activity appeared as discrete spherical sites, from < 0.3 to 1 μm in diameter, in the primary hyphae. In the secondary and tertiary branch hyphae of the older portions of the M. racemosus colony the sites were often as large as 3 μm in diameter. The sites of enzyme activity occurred sparsely and were randomly distributed in the hyphal tips. They were most numerous in older portions of these hyphae. Hydolysis of the p-nitrophenyl N-acetyl-β-d-glucosaminide was obtained with cell-free extracts of M. racemous, Rhizoctonia solani, Corticium sp., and Phycomyces nitens; however, cytochemical localizations of enzyme activity were observed only in M. racemosus: Four cytochemical substrates and 11 coupling agents were investigated. Only α-naphthyl N-acetyl-β-d-glucosaminide was satisfactory as a substrate. Fast Red B and p-nitrobenzene diazonium tetrafluoroborate were the only coupling reagents that yielded stable discrete reaction products. © 1979 Academic Press, Inc. All rights reserved.