title: Characterization of Cln3p, the gene product responsible for juvenile neuronal ceroid lipofuscinosis, as a lysosomal integral membrane glycoprotein.
abstract: Juvenile neuronal ceroid lipofuscinosis (JNCL) is an autosomal recessively inherited lysosomal storage disease involving a mutation in the CLN3 gene. The sequence of CLN3 was determined in 1995; however, the localization of the CLN3 gene product (Cln3p) was not confirmed. In this study, we investigated endogenous Cln3p using two peptide antibodies raised against two distinct epitopes of murine Cln3p. Identification of the liver 60 kDa protein as Cln3p was ascertained by amino acid sequence analysis using tandem mass spectrometry. Liver Cln3p was predominantly localized in the lysosomal membranes, not in endoplasmic reticulum (ER) or Golgi apparatus. As the tissue concentration of brain Cln3p was much lower than that of liver Cln3p, it could be detected only after purification from brain extract using anti-Cln3p IgG Sepharose. The apparent molecular masses of liver Cln3p and brain Cln3p were determined to be about 60 kDa and 55 kDa, respectively. Both brain and liver Cln3p were deglycosylated by PNGase F treatment to form polypeptides with almost the same molecular mass (45 kDa). However, they were not affected by Endo h treatment. In addition, it was also elucidated that the amino terminal region of Cln3p faces the cytosol.
authors: Department of Biochemistry, Central Laboratory of Medical Science, Juntendo University School of Medicine, Hongo 2-1-1, Bunkyo-ku, Tokyo 113, Japan.
authors: Junji
authors: J
authors: Ezaki
authors: Mitsue
authors: M
authors: Takeda-Ezaki
authors: Masato
authors: M
authors: Koike
authors: Yoshiyuki
authors: Y
authors: Ohsawa
authors: Hikari
authors: H
authors: Taka
authors: Reiko
authors: R
authors: Mineki
authors: Kimie
authors: K
authors: Murayama
authors: Yasuo
authors: Y
authors: Uchiyama
authors: Takashi
authors: T
authors: Ueno
authors: Eiki
authors: E
authors: Kominami
mesh_terms: D000818:Animals; D001923:Brain Chemistry; D002460:Cell Line; D006023:Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007163:Immunosorbent Techniques; D007425:Intracellular Membranes / Q000737:chemistry; D007668:Kidney / Q000737:chemistry / Q000166:cytology / Q000378:metabolism; D008099:Liver / Q000737:chemistry; D013058:Mass Spectrometry; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D051379:Mice; D018832:Molecular Chaperones; D008970:Molecular Weight; D009472:Neuronal Ceroid-Lipofuscinoses / Q000209:etiology*; D009844:Oligosaccharides / Q000737:chemistry; D009928:Organ Specificity; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013347:Subcellular Fractions / Q000737:chemistry
chemical_list: C102229:CLN3 protein, mouse; D006023:Glycoproteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D018832:Molecular Chaperones; D009844:Oligosaccharides; D011506:Proteins; C018589:lysosomal proteins