Phosphorylation and subcellular location of alpha-L-fucosidase in lymphoid cells from patients with I-cell disease and pseudo-Hurler polydystrophy.
Mannose 6-phosphate is a recognition marker used by many newly made acid hydrolases for their transport to lysosomes. Previously, we investigated the incorporation of 32Pi into alpha-L-fucosidase of lymphoid cell lines from a healthy individual (control) and an I-cell disease patient [DiCioccio and Miller, Glycobiology, 1, 595-604 (1991)]. Phosphoserine was found in immunoprecipitable alpha-L-fucosidase of both control and I-cell lymphoid cells, but mannose 6-phosphate was identified only in enzyme of control cells. Extension of this investigation to lymphoid cultures of a pseudo-Hurler polydystrophy patient also identified only phosphoserine in alpha-L-fucosidase. Using [3H]mannose instead of 32Pi, the precise identification of mannose 6-phosphate in alpha-L-fucosidase of control cells, and its absence in alpha-L-fucosidase of I-cell and pseudo-Hurler cells, was established. The stoichiometry of phosphorylation of alpha-L-fucosidase in I-cell, pseudo-Hurler and control lymphoid cells was 3, 4 and 10 mol Pi/mol enzyme, respectively. alpha-L-Fucosidase was located in lysosomes isolated from control, I-cell and pseudo-Hurler lymphoid cells by subcellular fractionation on Percoll density gradients. Both I-cell and pseudo-Hurler lymphoid cells displayed normal intralysosomal activity of alpha-L-fucosidase despite lack of the mannose 6-phosphate marker. Thus, I-cell and pseudo-Hurler lymphoid cells must possess a mannose 6-phosphate-independent mechanism for directing alpha-L-fucosidase to lysosomes.(ABSTRACT TRUNCATED AT 250 WORDS)
DiCioccio
R A
RA
Department of Gynecologic Oncology, Roswell Park Cancer Institute, Buffalo, NY 14263.
Miller
A L
AL
D002478:Cells, Cultured; D006801:Humans; D008214:Lymphocytes / Q000378:metabolism; D008358:Mannose / Q000378:metabolism; D008360:Mannosephosphates / Q000378:metabolism; D009081:Mucolipidoses / Q000201:enzymology* / Q000378:metabolism; D010766:Phosphorylation; D013347:Subcellular Fractions / Q000201:enzymology; D005644:alpha-L-Fucosidase / Q000737:chemistry / Q000378:metabolism*
D008360:Mannosephosphates; C027693:mannose-6-phosphate; D005644:alpha-L-Fucosidase; D008358:Mannose