An enzyme with properties similar to those of <i>β</i>-<i>N</i>-acetylhexosaminidase S is expressed in the promyelocytic cell line HL-60

Emiliani, Carla; Beccari, Tommaso; Tabilio, Antonio; Orlacchio, Aldo; Hosseini, Ramine; Stirling, J. L.

doi:10.1042/bj2670111

Cited by 26 publications

(21 citation statements)

References 22 publications

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“…In the brain extract from a Hexb À/À mouse (Figure 2b), only the Hex S isoenzyme (aa structure) was detected, eluting within the NaCl gradient just after the elution point of Hex A. 15 Treatment of Hexb À/À mice with Ad.HEXB resulted in the appearance of Hex B and Hex A isoforms in both the injected and contralateral hemispheres (Figure 2c and d, respectively). The restoration of Hex B activity reached a very high level, but the increase in Hex A was much weaker.…”

Section: Resultsmentioning

confidence: 99%

“…Brain extracts were analyzed by ion-exchange chromatography on DE-52 DEAE-cellulose (Whatmann Biochemicals, UK). 15 The chromatography was performed using 1 ml column equilibrated with 10 mM sodium phosphate buffer, pH 6.0. Proteins retained by the column were eluted using a linear gradient of NaCl (0.0-0.3 M in 30 ml of the above buffer).…”

Section: In Vivo Adhexb Administration In Hexb à/à Micementioning

confidence: 99%

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Widespread distribution of β-hexosaminidase activity in the brain of a Sandhoff mouse model after coinjection of adenoviral vector and mannitol

Bourgoin

Emiliani²,

Kremer³

et al. 2003

Gene Ther

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Sandhoff disease is a severe inherited neurodegenerative disorder resulting from deficiency of the b-subunit of hexosaminidases A and B, lysosomal hydrolases involved in the degradation of G M2 ganglioside and related metabolites. Currently, there is no viable treatment for the disease. Here, we show that adenovirus-mediated transfer of the bsubunit of b-hexosaminidase restored Hex A and Hex B activity after infection of Sandhoff fibroblasts. Gene transfer following intracerebral injection in a murine model of Sandhoff disease resulted in near-normal level of enzymatic activity in the entire brain at the different doses tested. The addition of hyperosmotic concentrations of mannitol to the adenoviral vector resulted in an enhancement of vector diffusion in the injected hemisphere. Adenoviral-induced lesions were found in brains injected with a high dose of the vector, but were not detected in brains injected with 100-fold lower doses, even in the presence of mannitol. Our data underline the advantage of the adjunction of mannitol to low doses of the adenoviral vector, allowing a high and diffuse transduction efficiency without viral cytotoxicity.

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Section: Resultsmentioning

confidence: 99%

Section: In Vivo Adhexb Administration In Hexb à/à Micementioning

confidence: 99%

Widespread distribution of β-hexosaminidase activity in the brain of a Sandhoff mouse model after coinjection of adenoviral vector and mannitol

Bourgoin

Emiliani²,

Kremer³

et al. 2003

Gene Ther

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“…Total Hex and Hex A activity was measured using 3 mM MUG and 3 mM MUGS, respectively, in 0.1 M citric acid/0.2 M disodium phosphate buffer, pH 4.5 [28].…”

Section: Determination Of Enzyme Activity and Protein Concentrationmentioning

confidence: 99%

“…The recovered supernatants were analyzed by ion-exchange chromatography on DE-52 DEAE-cellulose as previously described [28]. Briefly, a 1 ml column was equilibrated with 10 mM Na/P buffer, pH 6.0; protein retained by the column was eluted using a linear 0.0-0.5 M gradient of NaCl in 50 ml of the above buffer.…”

Section: Deae-chromatographymentioning

confidence: 99%

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Identification and characterization of mature β-hexosaminidases associated with human placenta lysosomal membrane

et al. 2008

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Hex (β-hexosaminidase) is a soluble glycohydrolase involved in glycoconjugate degradation in lysosomes, however its localization has also been described in the cytosol and PM (plasma membrane). We previously demonstrated that Hex associated with human fibroblast PM as the mature form, which is functionally active towards GM2 ganglioside. In the present study, Hex was analysed in a lysosomal membrane-enriched fraction obtained by purification from highly purified human placenta lysosomes. These results demonstrate the presence of mature Hex associated with the lysosomal membrane and displaying, as observed for the PM-associated form, an acidic optimum pH. When subjected to sodium carbonate extraction, the enzyme behaved as a peripheral membrane protein, whereas Triton X-114 phase separation confirmed its partially hydrophilic nature, characteristics which are shared with the PM-associated form of Hex. Moreover, two-dimensional electrophoresis indicated a slight difference in the pI of β-subunits in the membrane and the soluble forms of the lysosomal Hex. These results reveal a new aspect of Hex biology and suggest that a fully processed membrane-associated form of Hex is translocated from the lysosomal membrane to the PM by an as yet unknown mechanism. We present a testable hypothesis that, at the cell surface, Hex changes the composition of glycoconjugates that are known to be involved in intercellular communication and signalling.

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Platelet glycohydrolase activities: Characterization and release

Emiliani

Martino

Orlacchio

et al. 1995

Cell Biochemistry & Function

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Granules containing acid hydrolases have been detected in human platelets but have not been thoroughly characterized. We have studied the activity and characteristics of glycohydrolases present in normal human platelets, evaluated their release upon stimulation with thrombin, and assessed the contribution of platelet - released lysosomal contents to the glycohydrolase activity present in normal serum. Platelets contained a remarkable glycohydrolase activity with a prevalence of beta - N-acetylhexosaminidase. All glycohydrolases were released to some extent upon stimulation with thrombin and contributed to the glycohydrolase activity found in human serum. alpha-Mannosidase and alpha-galactosidase were partially inactivated after release by a mechanism as yet undefined. In addition, thrombin stimulation affects the intraplatelet isoenzyme pattern of beta-N-acetylhexosaminidase by producing the appearance of a new form.

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An enzyme with properties similar to those of β-N-acetylhexosaminidase S is expressed in the promyelocytic cell line HL-60

Cited by 26 publications

References 22 publications

Widespread distribution of β-hexosaminidase activity in the brain of a Sandhoff mouse model after coinjection of adenoviral vector and mannitol

Widespread distribution of β-hexosaminidase activity in the brain of a Sandhoff mouse model after coinjection of adenoviral vector and mannitol

Identification and characterization of mature β-hexosaminidases associated with human placenta lysosomal membrane

Platelet glycohydrolase activities: Characterization and release

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