A lysosomal cysteine proteinase from dictyostelium discoideum contains N-acetylglucosamine-1-phosphate bound to serine but not mannose-6-phosphate on N-linked oligosaccharides.

Previous studies showed that vegetative Dictyostelium
discoideum cells make a lysosomal proteinase, proteinase-
1, that contains multiple GlcNAc-a-1-P residues
in phosphodiester linkage to serine. We extended these
studies and, in contrast to earlier reports, found that
proteinase-1 contains 7.5 mol of Fuc, 8 mol of Man, 2 mol
of Xyl, and 30 mol of GlcNAc per calculated mol of protein
but no Man-6-P residues. The protein binds to concanavalin
A and wheat germ agglutinin lectin affinity
columns, and PNGase-F digestion released most of the
mannose and xylose but little of the GlcNAc. b-Elimination
under reducing conditions released only GlcNAc-a-
1-P. There was no evidence for the release of disaccharides
or of fucitol. A rabbit antiserum and monoclonal
antibodies prepared against proteinase-1 recognize
GlcNAc-a-1-P residues in immunoblots and are specifically
competed by UDP-GlcNAc or GlcNAc-a-1-P. Use of
other monoclonal antibodies showed the presence of
mannose-6-sulfate on N-linked sugar chains, and a-fucose
residues on the protein. Thus, proteinase-1 has at
least two types of modifications: GlcNAc-a-1-P-Ser,
which we call phosphoglycosylation, and N-linked oligosaccharides.
This is the first purified lysosomal enzyme
in Dictyostelium that does not contain Man-6-P residues.
The GlcNAc-a-1-P-specific antibodies also recognize a
group of developmentally regulated proteins, especially
enriched in vegetative cells. Some of them are also lysosomal
cysteine proteinases, and all bind to the GlcNAca-
1-P-specific monoclonal antibody but not to the mammalian
CI-Man-6-P receptor. Conversely, lysosomal
enzymes that have Man-6-P do not bind to the GlcNAca-
1-P-specific antibody. An exception to this is b-Nacetylglucosaminidase,
where 15% of the activity binds
to this antibody. Thus, there appear to be two sets of
lysosomal enzymes with distinct post-translational
modifications.