title: Molecular characterization of mutations that cause globoid cell leukodystrophy and pharmacological rescue using small molecule chemical chaperones.
abstract: Globoid cell leukodystrophy (GLD) (Krabbe disease) is an autosomal recessive, degenerative, lysosomal storage disease caused by a severe loss of galactocerebrosidase (GALC) enzymatic activity. Of the >70 disease-causing mutations in the GALC gene, most are located outside of the catalytic domain of the enzyme. To determine how GALC mutations impair enzymatic activity, we investigated the impact of multiple disease-causing mutations on GALC processing, localization, and enzymatic activity. Studies in mammalian cells revealed dramatic decreases in GALC activity and a lack of appropriate protein processing into an N-terminal GALC fragment for each of the mutants examined. Consistent with this, we observed significantly less GALC localized to the lysosome and impairment in either the secretion or reuptake of mutant GALC. Notably, the D528N mutation was found to induce hyperglycosylation and protein misfolding. Reversal of these conditions resulted in an increase in proper processing and GALC activity, suggesting that glycosylation may play a critical role in the disease process in patients with this mutation. Recent studies have shown that enzyme inhibitors can sometimes "chaperone" misfolded polypeptides to their appropriate target organelle, bypassing the normal cellular quality control machinery and resulting in enhanced activity. To determine whether this may also work for GLD, we examined the effect of alpha-lobeline, an inhibitor of GALC, on D528N mutant cells. After treatment, GALC activity was significantly increased. This study suggests that mutations in GALC can cause GLD by impairing protein processing and/or folding and that pharmacological chaperones may be potential therapeutic agents for patients carrying certain mutations.
authors.affiliation: Department of Neuroscience, Mayo Clinic, Jacksonville, Florida 32224, USA.
authors.forename: Wing C
authors.initials: WC
authors.lastname: Lee
authors.forename: Dongcheul
authors.initials: D
authors.lastname: Kang
authors.forename: Ena
authors.initials: E
authors.lastname: Causevic
authors.forename: Aimee R
authors.initials: AR
authors.lastname: Herdt
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Eckman
authors.forename: Christopher B
authors.initials: CB
authors.lastname: Eckman
mesh_terms: D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D004789:Enzyme Activation / Q000187:drug effects / Q000235:genetics; D004792:Enzyme Precursors / Q000235:genetics / Q000378:metabolism; D005698:Galactosylceramidase / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007965:Leukodystrophy, Globoid Cell / Q000188:drug therapy* / Q000201:enzymology / Q000235:genetics*; D018832:Molecular Chaperones / Q000235:genetics* / Q000494:pharmacology / Q000627:therapeutic use*; D016297:Mutagenesis, Site-Directed; D017510:Protein Folding / Q000187:drug effects; D011499:Protein Processing, Post-Translational / Q000187:drug effects / Q000235:genetics
chemical_list: D004792:Enzyme Precursors; D018832:Molecular Chaperones; D005698:Galactosylceramidase
references.citation: J Biol Chem. 1988 Jan 25;263(3):1424-31
references.pmid: 3257215
references.citation: J Clin Invest. 1989 May;83(5):1590-7
references.pmid: 2496144
references.citation: Hum Mol Genet. 1993 Nov;2(11):1841-5
references.pmid: 8281145
references.citation: J Hum Genet. 2006;51(6):548-554
references.pmid: 16607461
references.citation: Nature. 1992 Aug 27;358(6389):761-4
references.pmid: 1380673
references.citation: Exp Eye Res. 2006 Jun;82(6):1030-6
references.pmid: 16297911
references.citation: Biochem Biophys Res Commun. 1994 Jan 28;198(2):485-91
references.pmid: 8297359
references.citation: Eur J Neurosci. 1995 Jun 1;7(6):1245-65
references.pmid: 7582098
references.citation: J Inherit Metab Dis. 1999 Apr;22(2):155-62
references.pmid: 10234611
references.citation: Biochim Biophys Acta. 1993 Sep 29;1170(1):53-61
references.pmid: 8399327
references.citation: Hum Mutat. 2007 Jul;28(7):742
references.pmid: 17579360
references.citation: FASEB J. 2005 Sep;19(11):1549-51
references.pmid: 15987783
references.citation: Clin Chim Acta. 1977 May 16;77(1):53-9
references.pmid: 17489
references.citation: Nat Genet. 2005 Jul;37(7):692-700
references.pmid: 15924140
references.citation: Clin Chim Acta. 1998 Aug 10;276(1):53-61
references.pmid: 9760019
references.citation: Am J Pathol. 1988 May;131(2):308-19
references.pmid: 3358457
references.citation: J Neurosci. 2007 Dec 12;27(50):13730-8
references.pmid: 18077684
references.citation: FEBS J. 2007 Oct;274(19):4962-71
references.pmid: 17894781
references.citation: Thromb Haemost. 2001 Oct;86(4):1023-7
references.pmid: 11686319
references.citation: Brain Res. 1988 Jun 28;454(1-2):340-6
references.pmid: 3409017
references.citation: Curr Opin Chem Biol. 2007 Aug;11(4):412-8
references.pmid: 17644022
references.citation: Genomics. 1996 Sep 15;36(3):468-75
references.pmid: 8884270
references.citation: Proc Natl Acad Sci U S A. 1970 Jun;66(2):302-9
references.pmid: 5271165
references.citation: Hum Genet. 1997 Sep;100(3-4):450-6
references.pmid: 9272171
references.citation: Hum Mutat. 1997;10(4):268-79
references.pmid: 9338580
references.citation: J Neurosci. 2009 May 13;29(19):6068-77
references.pmid: 19439584
references.citation: J Biol Chem. 2000 Apr 21;275(16):12281-9
references.pmid: 10766867
references.citation: Arch Neurol. 1999 Aug;56(8):1014-7
references.pmid: 10448809
references.citation: Hum Genet. 1996 Mar;97(3):304-8
references.pmid: 8786069
references.citation: Am J Hum Genet. 1996 Dec;59(6):1233-42
references.pmid: 8940268
references.citation: Mol Genet Metab. 2000 May;70(1):1-9
references.pmid: 10833326
references.citation: J Neurol. 2000 Nov;247(11):875-7
references.pmid: 11151421
references.citation: Chem Biol. 2005 Nov;12(11):1235-44
references.pmid: 16298303
references.citation: J Mol Neurosci. 2002 Aug-Oct;19(1-2):129-33
references.pmid: 12212770
references.citation: Biol Chem. 2008 Jan;389(1):1-11
references.pmid: 18095864
references.citation: Brain Res. 1986 Apr;391(1):79-84
references.pmid: 3513905
references.citation: Bol Estud Med Biol. 1981 Jul-Sep;31(7-8):373-82
references.pmid: 7347616