Expression and processing of recombinant human galactosylceramidase

Nagano, Sukehisa; Yamada, Takeshi; Shinnoh, Nobue; Furuya, Hirokazu; Taniwaki, Takayuki; Kira, Jun Ichi

doi:10.1016/s0009-8981(98)00095-3

Cited by 39 publications

(32 citation statements)

References 14 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…16 Finally we examined the expression pattern of hGALC in the brain of the rescued animals using antibodies raised against a peptide contained in the 30 KDa subunit. 22 This analysis showed GALC staining of cells throughout the brain of the rescued mouse. An example of cerebellar staining is shown in Figure 2b where both cells in the white matter as well as Purkinje cell neurons contain reaction product.…”

Section: Expression Of the Human Galc Transgenementioning

confidence: 89%

“…Polyclonal antibodies recognizing the 30 kDa mature GALC subunit were raised in rabbits using the synthetic peptide CTGRKGSYPLPPKSQPF (spanning amino acids 455-469 of the human GALC; modified from Nagano et al 22 ) conjugated to keyhole limpet hemocyanin (CTG Transgenic rescue of Krabbe disease R De Gasperi et al peptide). The antibodies were subsequently purified by affinity chromatography through CTG-agarose.…”

Section: Preparation Of Antibodies Against Galcmentioning

confidence: 99%

See 1 more Smart Citation

Transgenic rescue of Krabbe disease in the twitcher mouse

et al. 2004

View full text Add to dashboard Cite

Dedicated with respect and appreciation to Professor Robert A Lazzarini, our teacher and mentor, who made all this research possibleThe twitcher mouse is a natural model of Krabbe disease caused by galactocerebrosidase (GALC) deficiency. Previous attempts at rescuing the twitcher mouse by bone marrow transplantion, viral transduction, or transgenesis were only partially successful. Here, we report the transgenic (tg) rescue of the twitcher mouse with a BAC clone harboring the entire GALC. The twi/twi/hGALC tg mice exhibited growth, motor function, and fertility similar to those of nonaffected animals. These animals had normal levels of GALC activity in brain and were free of the typical twitcher demyelinating pathology. Surprisingly, GALC expression in twi/twi hGALC tg kidneys was low and galactocerebroside storage was only partially cleared. Nonetheless, these mice have been maintained for over 1 year without any sign of disease. Since pathological damage associated with GALC deficiency is confined to the nervous system, our work represents the first successful rescue of the twitcher mouse and opens the possibility of developing novel therapeutic approaches.

show abstract

Section: Expression Of the Human Galc Transgenementioning

confidence: 89%

Section: Preparation Of Antibodies Against Galcmentioning

confidence: 99%

Transgenic rescue of Krabbe disease in the twitcher mouse

et al. 2004

View full text Add to dashboard Cite

show abstract

“…[1] After entering the lysosomes, the enzyme is cleaved into 30-and 50-kDa subunits without effect on enzymatic activity. [2] Crystal studies indicate that no dissociation of these subunits occurs. [3] Substrate hydrolysis by GALC occurs through a Koshland double displacement mechanism with overall retention of the β-anomeric configuration of the released galactopyranoside ( Figure 1A).…”

Section: Introductionmentioning

confidence: 99%

“…The mechanism behind this neuropathology has not yet been fully elucidated. [1,2] Reduced activity of GALC results in a reduced catabolism of galactosphingolipids including galactosylceramide and galactosylsphingosine (psychosine). Galactosylceramide is the main lipid component of myelin, the protective sheath around neuron axons and essential for correct functioning of the nervous system.…”

Section: Introductionmentioning

confidence: 99%

A Specific Activity‐Based Probe to Monitor Family GH59 Galactosylceramidase, the Enzyme Deficient in Krabbe Disease

et al. 2017

View full text Add to dashboard Cite

Galactosylceramidase (GALC) is the lysosomal β-galactosidase responsible for the hydrolysis of galactosylceramide. Inherited deficiency in GALC causes Krabbe disease, a devastating neurological disorder characterized by accumulation of galactosylceramide and its deacylated counterpart, the toxic sphingoid base galactosylsphingosine (psychosine). We report the design and application of a fluorescently tagged activity-based probe (ABP) for the sensitive and specific labeling of active GALC molecules from various species. The probe consists of a β-galactopyranose-configured cyclophellitol-epoxide core, conferring specificity for GALC, equipped with a Bodipy fluorophore at C6 that allows visualizing active enzyme in cells and tissues. The detection of residual GALC in patient fibroblasts holds great promise for laboratory diagnosis of Krabbe disease. We further describe a procedure for in situ imaging of active GALC in murine brain by intracerebroventricular infusion of the ABP. In conclusion, the GALCspecific ABP should find broad applications in diagnosis, drug development and evaluation of therapy of Krabbe disease.

show abstract

“…1) that is unlikely to harbor hydrolase activity because it lacks corresponding catalytic residues (8). The three domains pack against one another with high buried surface areas; however, in cell culture, 53-and 30-kDa cleavage adducts have been detected (10), suggesting the lectin domain may be more mobile and, as a consequence, susceptible to proteolysis in solution. The evolutionary origin and function of the lectin domain will likely be the subject of future work in the field.…”

mentioning

confidence: 99%

Sneak peak at galactocerebrosidase, Krabbe disease's lysosomal hydrolase

Lieberman

2011

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Expression and processing of recombinant human galactosylceramidase

Cited by 39 publications

References 14 publications

Transgenic rescue of Krabbe disease in the twitcher mouse

Transgenic rescue of Krabbe disease in the twitcher mouse

A Specific Activity‐Based Probe to Monitor Family GH59 Galactosylceramidase, the Enzyme Deficient in Krabbe Disease

Sneak peak at galactocerebrosidase, Krabbe disease's lysosomal hydrolase

Contact Info

Product

Resources

About