Lysosomal enzyme targeting. N-Acetylglucosaminylphosphotransferase selectively phosphorylates native lysosomal enzymes.
Lysosomal enzymes contain 6-phosphomannosyl moieties which mediate their translocation to lysosomes. This recognition marker is synthesized by the sequential action of UDP-N-acetylglucosamine:lysosomal enzyme N-acetylglucosamine-1-phosphotransferase and alpha-N-acetylglucosaminyl phosphodiesterase. A new assay for the N-acetylglucosaminylphosphotransferase, using alpha-methylmannoside as acceptor, is presented. Using this assay, we partially purified the transferase and examined its substrate specificity. The transferase exhibited a very high affinity toward lysosomal enzymes (apparent Km values of less than 20 microM) and was greater than 100-fold more efficient (Vmax/Km) when using lysosomal enzymes as acceptors as compared to nonlysosomal glycoproteins that contain high mannose oligosaccharide units. Heat denaturation of the lysosomal enzymes resulted in the loss of acceptor activity. The model compounds alpha-methylmannoside and Man5--8GlcNAc were poor acceptors. We propose that this enzyme catalyzes the initial, determining step by which synthesized acid hydrolases are distinguished from other newly synthesized glycoproteins and thus are eventually targeted to lysosomes.
D000118:Acetylglucosaminidase / Q000378:metabolism; D000818:Animals; D004288:Dolichol Phosphates / Q000378:metabolism; D005964:Glucosyltransferases / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008247:Lysosomes / Q000201:enzymology*; D009844:Oligosaccharides; D010766:Phosphorylation; D051381:Rats; D011919:Rats, Inbred Strains; D013379:Substrate Specificity; D017855:Transferases (Other Substituted Phosphate Groups)
D004288:Dolichol Phosphates; D009844:Oligosaccharides; D005964:Glucosyltransferases; D017855:Transferases (Other Substituted Phosphate Groups); C023222:UDPacetylglucosamine-dolichyl-phosphate acetylglucosamine-1-phosphate transferase; D000118:Acetylglucosaminidase