Studies on human β D N acetylhexosaminidases. I. Purification and properties

Satish Srivastava, Y. C. Awasthi, A. Yoshida, E. Beutler

Research output: Contribution to journalArticle

79 Citations (Scopus)

Abstract

β (D) N Acetylhexosaminidase A and B were purified from human placenta. By extraction, ammonium sulfate precipitation, lyophilization, Sephadex G 200 filtration, and DEAE cellulose column chromatography, hexosaminidase A and hexosaminidase B were purified together. Hexosaminidase A was further purified by a second DEAE cellulose column chromatography followed by ECTEOLA cellulose column chromatography, Sephadex G 200 filtration, electrofocusing, and a third Sephadex G 200 filtration. Hexosaminidase B was further purified by CM cellulose column chromatography, electrofocusing, and Sephadex G 200 filtration. After the first Sephadex G 200 filtration hexosaminidase A and B were also purified by a second method. This method involved calcium phosphate gel, DEAE Sephadex, ECTEOLA cellulose, CM Sephadex and CM cellulose column chromatography, and preparative polyacrylamide disc electrophoresis. Purified hexosaminidase A and hexosaminidase B moved on polyacrylamide disc electrophoresis as single protein bands with virtually all of the enzyme activity. Hexosaminidase A and B in 10 mM phosphate buffer containing 0.1 M (NH4)2SO4 were stable at 4° for at least 4 mth. The isoelectric points of hexosaminidase A and hexosaminidase B were found to be pH 5.4 and 7.9, respectively. Placental hexosaminidases differed electrophoretically and antigenically from the liver and fibroblast enzymes. In contrast to the reports of others neuraminidase failed to convert heat labile hexosaminidase A to a heat stable form of enzyme resembling hexosaminidase B.

Original languageEnglish (US)
Pages (from-to)2043-2048
Number of pages6
JournalJournal of Biological Chemistry
Volume249
Issue number7
StatePublished - 1974
Externally publishedYes

Fingerprint

Hexosaminidase B
Hexosaminidase A
Purification
Column chromatography
Chromatography
Disc Electrophoresis
DEAE-Cellulose Chromatography
DEAE-Cellulose
Isoelectric Focusing
Electrophoresis
Cellulose
Enzymes
Hot Temperature
Hexosaminidases
DEAE-Dextran
Freeze Drying
Isoelectric Point
Ammonium Sulfate
Neuraminidase
Enzyme activity

ASJC Scopus subject areas

  • Biochemistry

Cite this

Srivastava, S., Awasthi, Y. C., Yoshida, A., & Beutler, E. (1974). Studies on human β D N acetylhexosaminidases. I. Purification and properties. Journal of Biological Chemistry, 249(7), 2043-2048.

Studies on human β D N acetylhexosaminidases. I. Purification and properties. / Srivastava, Satish; Awasthi, Y. C.; Yoshida, A.; Beutler, E.

In: Journal of Biological Chemistry, Vol. 249, No. 7, 1974, p. 2043-2048.

Research output: Contribution to journalArticle

Srivastava, S, Awasthi, YC, Yoshida, A & Beutler, E 1974, 'Studies on human β D N acetylhexosaminidases. I. Purification and properties', Journal of Biological Chemistry, vol. 249, no. 7, pp. 2043-2048.
Srivastava S, Awasthi YC, Yoshida A, Beutler E. Studies on human β D N acetylhexosaminidases. I. Purification and properties. Journal of Biological Chemistry. 1974;249(7):2043-2048.
Srivastava, Satish ; Awasthi, Y. C. ; Yoshida, A. ; Beutler, E. / Studies on human β D N acetylhexosaminidases. I. Purification and properties. In: Journal of Biological Chemistry. 1974 ; Vol. 249, No. 7. pp. 2043-2048.
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