Purification and physical-chemical characterization of hepatic stimulator substance

Authors

  • Dr. Douglas R. Labrecque M.D.,

    Director, Corresponding author
    1. Iowa City Veterans Administration Hospital and the Liver Service, University of Iowa Center for Digestive Diseases Iowa City, Iowa 52242
    • Liver Service, Center for Digestive Diseases, University of Iowa Hospitals and Clinics, Iowa City, Iowa 52242
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    • Dr. LaBrecque performed some of these studies while a Clinical Investigator at the Iowa City Veterans Administration Medical Center.

  • Gregory Steele,

    1. Iowa City Veterans Administration Hospital and the Liver Service, University of Iowa Center for Digestive Diseases Iowa City, Iowa 52242
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  • Steven Fogerty,

    1. Iowa City Veterans Administration Hospital and the Liver Service, University of Iowa Center for Digestive Diseases Iowa City, Iowa 52242
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  • Michelle Wilson,

    1. Iowa City Veterans Administration Hospital and the Liver Service, University of Iowa Center for Digestive Diseases Iowa City, Iowa 52242
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  • James Barton

    1. Iowa City Veterans Administration Hospital and the Liver Service, University of Iowa Center for Digestive Diseases Iowa City, Iowa 52242
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  • This study is dedicated to the memory of Dr. Edward Heath, Chief, Department of Biochemistry, University of Iowa, 1976–1984.

Abstract

Hepatic stimulator substance is a liver growth stimulator derived from the hepatocyte cytosol of weanling or regenerating adult rat livers. The present paper reports the almost 9,000-fold purification of hepatic stimulator substance with an approximately 100,000-fold increase in specific growth stimulator activity. Purification steps included heating at 95°C for 15 min, 40% cold ethanol precipitation, passage over Procion Red HE3B, DEAE cellulose and Sephadex G75 columns and gel filtration and reverse-phase fast protein liquid chromatography techniques. As little as 27 ng per ml of the purest material produced a 2-fold stimulation in the standard HTC cell activity assay. Further studies indicate that hepatic stimulator substance is a highly negatively charged protein and that disulfide bonds or a complex tertiary structure are not essential to its activity. Hepatic stimulator substance is stable over a wide range of pH's and temperatures. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis with silver stain revealed 1 major band at 12,400 daltons and 1 minor band at 17,500 daltons.

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