Identification of tay-sachs disease carriers by acrylamide gel electrophoresis

Friedland, Joan; Schneck, Larry; Saifer, Abraham; Pourfar, M.; Volk, Bruno W.

doi:10.1016/0009-8981(70)90064-1

Cited by 53 publications

(11 citation statements)

References 11 publications

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“…kidney (6), spleen (5), and leukocytes (8). Both fractions, A and B, were examined for P-D-glucosidase, P-D-galactosidase, and acid phosphatase activity, but none of these activities could be detected.…”

Section: Enzymatic Properties Of Enzyme Forms a And Bmentioning

confidence: 99%

The Partial Purification of Two β-N-Acetyl-D-hexosaminidases from Porcine Kidney

Wetmore¹,

Verpoorte²

1972

Can. J. Biochem.

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WETMORE, S. J., and VERPBORTE, J. A. The partial purification of two B-N-acetyl-D-hexosaminidases from porcine kidney. Can. J. Biochem. 50,563-573 (1972).Two distinct fractions showing both B-N-acetyl-D-glucosaminidase (EC 3.2.1.30) and 8-N-acetyl-sgalactowminidase activity were isolated and purified from pig kidney. These preparations, which were designated A and B, were not stable during gel chromatography or prolonged dialysis. Final purifications of 600-fold for enzyme A and 440-fold for enzyme B were obtained.Gel electrophoresis and ultracentrifugation studies indicated heterogeneity in both preparations. The amino acid compositions of both preparations were very similar. Ultracentrifugation studies suggested the formation of subunits in the presence of 5 M guanidine-HCl and 1 mM dithiothreitol.A study of the enzymatic properties also showed great similarities between the two enzyme forms. Both enzymes had identical Michaelis-Menten constants of 1.88 mM forp-nitrophenyl-8-N-acetyl-D-glucosaminide and 0.38 mM for p-nitrophenyl-B-N-acetyl-D-galactosaminide. Although bovine serum albumin enhanced the activity of the enzymes it did not change the K, values. The pH-rate profiles of both enzymes with the substrate p-nitrophenyl-B-N-acetyl-D-glucodnide showed two peaks. When p-nitrophenyl-p-N-acetyl-Dgalactosaminide was used as substrate, only one peak was observed in the pH-rate profiles. However, in this case a distinct shoulder could be detected in these peaks. Heating at SO0 destroyed the activities of both forms of the enzyme rapidly, but addition of bovine senun albumin protected against heat inactivation. WETMORE, S. J., et VERPOBRTE, J. A. The partial purification of two B-N-acetyl-D-hexosaminidases from porcine kidney. Can. J. Biochem. 58, 563-573 (1972). Deux fractions distinctes poss&bnt chacune une B-N-acktyl-D-glucosaminidase (EC 3.2.1.30) et une B-N-acktyl-D-galactosaminidase sont isolbes et purifiks a partir du rein de porc. Ces dew preparations, nommtes A et B, sont instables durant la chromatographie sur gel ou durant une dialyse prolongee. La purification finale de l'enzyme A est de 600 fois et celle de l'enzyme B, de 440 fois. L'tlectrophorese sur gel et l'ultracentrifugation dtmontrent l'httkroginkitt des deux preparations. Les compositions en acides amints des deux preparations se ressemblent beaucoup. Les rbultats de l'ultracentrifugation laissent croire A la formation de sous-unites en presence de guanidine-HC1 5 M et de dithiothrkitol 1 mM. L'etude des proprietes enzymatiques rkvele aussi une grande similitude entre les deux enzymes. Tous dew ont la meme constante de Michaelis-Menten pour le p-nitrophknyl-B-N-acktyl-D-glucosaminide (1.88 mM) et pour le p-nitrophtnyl-B-N-acttyl-s-galactoaminde (0.38 mM). Bien que l'albumine krique du boeuf augmente l'activitk de ces enzymes, elle ne change par leurs valeurs de K,. Avec lep-nitrophknyl-8-N-acktyl-Dgucosaminide, les courbes en fonction du pH montrent deux sommets. Avec le p-nitrophknyl-B-N-acCtyl-Dgalactosaminide, ces courbes n'offrent plus qu'...

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Section: Enzymatic Properties Of Enzyme Forms a And Bmentioning

confidence: 99%

The Partial Purification of Two β-N-Acetyl-D-hexosaminidases from Porcine Kidney

Wetmore¹,

Verpoorte²

1972

Can. J. Biochem.

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“…The enzyme determination of hexosaminidase A and B activities in the brain, and liver amniotic cells and fluid was performed by acrylamide gel electrophoresis using 4-methylumbelliferyl substrate (8). The ganglioside extraction and chromatographic isolation procedures were carried out according to the methods of Folch et al (9) and Rouser et at?.…”

mentioning

confidence: 99%

Early Evolution Of Cytoplasmic Inclusion Bodies in Tay-Sachs Disease

Volk

Adachi

Friedland

et al. 1970

Experimental Biology and Medicine

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“…The lower anodic reservoir was filled with 0.1 mol/1 Tris-HCl buffer, pH 8.1. The upper cathodic buffer consisted of 4 g Tris and 7.2 g Tricine (Sigma) dissolved in a litre of distilled water at pH 8.1, as used by Friedtandet al (20). After setting the apparatus the glass cells were completely immersed in the buffer so that no excessive heating could be produced in the gel.…”

Section: Electrophoresismentioning

confidence: 99%

A Microelectrophoretic Method for the Separation of β-N-Acetylglucosaminidase A and B from Cultured Human Fibroblasts and Amniotic Cells with the Aid of Polyacrylamide Flat Disc Gels

Singh¹,

Hw²

1975

Clinical Chemistry and Laboratory Medicine

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Summary: A flat gel polyacrylamide microelectrophoretic method is described enabling the separation of human hexosaminidase A and B from cultured human fibroblasts and amniotic cells in parallel multiple channels. The utility of this method in prenatal diagnosis of Tay-Sachs and Sandhoff s disease has been emphasised. Eine mikroelektrophoretische Methode zur Trennung der ß-N-AcetylglucosaminidaseA und B von kultivierten Fibroblasten des Menschen und Amnionzellen mit Hilfe von Polyacrylamid-DiskflacligelenZusammenfassung: Eine Mikromethode zur Auftrennung der Hexosaminidäsen A und B aus Fibroblastenkulturen und Amnionzellen mittels Polyacrylamidgel-Elektrophorese wird beschrieben; sie ist für die pränatale Diagnose der Tay-Sachs-und Sa/jd/io/jf-Krankheit geeignet.

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Identification of tay-sachs disease carriers by acrylamide gel electrophoresis

Cited by 53 publications

References 11 publications

The Partial Purification of Two β-N-Acetyl-D-hexosaminidases from Porcine Kidney

The Partial Purification of Two β-N-Acetyl-D-hexosaminidases from Porcine Kidney

Early Evolution Of Cytoplasmic Inclusion Bodies in Tay-Sachs Disease

A Microelectrophoretic Method for the Separation of β-N-Acetylglucosaminidase A and B from Cultured Human Fibroblasts and Amniotic Cells with the Aid of Polyacrylamide Flat Disc Gels

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