PMID: 6539623Jul 6, 1984Paper

Purification and characterization of acetoacetyl-CoA synthetase from rat liver

Biochimica Et Biophysica Acta
M ItoK Tomita

Abstract

Acetoacetyl-CoA synthetase (acetoacetate:CoA ligase) was purified to electrophoretic homogeneity from a rat liver supernatant by ammonium sulfate fractionation and successive chromatographies on DEAE-Sepharose, Blue-Sepharose, Red-Sepharose, CoA-Sepharose, Ultrogel AcA-44, and DEAE-Sepharose once again. The purified enzyme had a specific activity of 2.3 mumol acetoacetyl-CoA formed per min per mg protein, which constituted a 960-fold purification compared to the crude extract, with a 7.4% yield. The enzyme absolutely required ATP, CoA, a monovalent cation (K+, Rb+, Cs+ or NH4+) and a divalent cation (Mg2+, Mn2+ or Ni2+) for its activity, yielding acetoacetyl-CoA, AMP and PPi in equimolar amounts. The molecular weight of the enzyme was approximately 80 000 as determined by Ultrogel AcA-34 gel filtration, and 71 000 by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The enzyme was active only on acetoacetate and to a lesser extent on L-(+)-3-hydroxybutyrate, and the Km values for acetoacetate, L-(+)-3-hydroxybutyrate, ATP and CoA were 8 microM, 75 microM, 60 microM and 10 microM, respectively.

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Citations

Oct 7, 2008·Applied Microbiology and Biotechnology·Hisashi HaradaNorihiko Misawa
May 30, 2002·Biochemical Pharmacology·Hiroki SatoTetsuya Fukui
Feb 12, 2013·American Journal of Physiology. Heart and Circulatory Physiology·David G CotterPeter A Crawford
Sep 25, 2012·Biochemical and Biophysical Research Communications·Shinya HasegawaTetsuya Fukui
Feb 17, 2009·Obesity·Ryota NarishimaTetsuya Fukui
Mar 30, 2016·Biochemical and Biophysical Research Communications·Masahiro YamasakiTetsuya Fukui
Nov 6, 2012·Biological & Pharmaceutical Bulletin·Shinya HasegawaTetsuya Fukui
Jan 5, 2011·Neuroscience Letters·Ryota NarishimaTetsuya Fukui
Oct 11, 2014·Biochemical and Biophysical Research Communications·Shinya HasegawaTetsuya Fukui
May 1, 2016·FEBS Letters·Shinya HasegawaTetsuya Fukui
Aug 2, 2005·Biochemical and Biophysical Research Communications·Masahiro YamasakiTetsuya Fukui
Nov 8, 2005·Journal of Biochemistry·Yoshinori KuboNoriko Takahashi

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