Enzyme product blot for nondestructive assay of protein catalytic function in polyacrylamide gels

Mikeanne Minter, Jeffrey Towbin, Jeffrey Harter, Edward R.B. McCabe

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

A new method that permits rapid, sensitive, and specific enzymatic assay of proteins in polyacrylamide gels is described. The enzyme product blot described in this report involves percolation of the reaction mixture through a gel containing native enzyme which converts the labeled substrate to a labeled product with differing chemical properties. A permeable membrane with specific ligand-binding properties overlies the gel and binds the enzyme product, but not the substrate, as reaction mixture is blotted vertically. This membrane is washed free of substrate and the location of the product is identified by autoradiography. The autoradiogram is compared with the stained gel in order to recover the enzyme for amino acid sequence analysis. The enzyme product blot is demonstrated using glycerol kinase and hexokinase.

Original languageEnglish (US)
Pages (from-to)22-26
Number of pages5
JournalAnalytical Biochemistry
Volume178
Issue number1
DOIs
StatePublished - Jan 1 1989
Externally publishedYes

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Assays
Enzymes
Gels
Proteins
Substrates
Glycerol Kinase
Membranes
Hexokinase
Protein Sequence Analysis
Enzyme Assays
Autoradiography
Chemical properties
polyacrylamide gels
Ligands
Amino Acids

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Enzyme product blot for nondestructive assay of protein catalytic function in polyacrylamide gels. / Minter, Mikeanne; Towbin, Jeffrey; Harter, Jeffrey; McCabe, Edward R.B.

In: Analytical Biochemistry, Vol. 178, No. 1, 01.01.1989, p. 22-26.

Research output: Contribution to journalArticle

Minter, Mikeanne ; Towbin, Jeffrey ; Harter, Jeffrey ; McCabe, Edward R.B. / Enzyme product blot for nondestructive assay of protein catalytic function in polyacrylamide gels. In: Analytical Biochemistry. 1989 ; Vol. 178, No. 1. pp. 22-26.
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