RAT-BRAIN CONTAINS HIGH-LEVELS OF MANNOSE-6-PHOSPHORYLATED GLYCOPROTEINS INCLUDING LYSOSOMAL-ENZYMES AND PALMITOYL-PROTEIN THIOESTERASE, ANENZYME IMPLICATED IN INFANTILE NEURONAL LIPOFUSCINOSIS

Mannose 6-phosphate (Man-B-P) is a posttranslational carbohydrate modi fication typical of newly synthesized acid hydrolases that signals tar geting from the Gels apparatus to the lysosome via Man-B-P receptors ( MPRs), Using iodinated cation independent MPR as a probe in a Western blot assay, we surveyed levels of Man-B-P glycoproteins in a number of different rat tissues, Considerable variation was observed with respe ct to total amounts and types of Man-B-P glycoproteins in the differen t tissues, Brain contained 2-8-fold more Man-B-P glycoproteins than ot her tissues, with relative abundance being brain much greater than tes tis approximate to heart > lung approximate to kidney approximate to o vary approximate to spleen > skeletal muscle approximate to liver appr oximate to serum, Analysis of 16 different lysosomal enzyme activities revealed that brain contains lower activities than other tissues whic h suggested that decreased removal of Man-6-P results in increased lev els of Man-B-P glycoproteins, This was directly demonstrated by compar ing activities of phosphorylated lysosomal enzymes, purified by im mob ilized MPR affinity chromatography, with total activities, The phospho rylated forms accounted for a considerable proportion of the MPR-targe ted activities measured in brain (on average, 36.2%) but very little i n lung, kidney, and liver (on average, 5.5, 2.3, and 0.7%, respectivel y), Man-B-P glycoproteins were also isolated from rat brain by MPR aff inity chromatography on a preparative scale. Of the 18 bands resolvabl e by SDS-polyacrylamide gel electrophoresis, seven bands were NH2-term inally sequenced and identified as the known lysosomal enzymes catheps in L, cathepsin A, cathepsin D, alpha-galactosidase A, arylsulfatase A , and alpha-iduronidase, One of the major Man-6-P glycoproteins was id entified as palmitoyl protein thioesterase, which was not previously t hought to be lysosomal, This finding raises important questions about the cellular location and function of palmitoyl protein thioesterase, mutations in which result in the neurodegenerative disorder, infantile neuronal ceroid lipofuscinosis.