PMID: 41873Oct 1, 1979

Chemical modification of horseradish peroxidase. Preparation and characterization of tracer enzymes with different isoelectric points

The Journal of Histochemistry and Cytochemistry : Official Journal of the Histochemistry Society
H G Rennke, M A Venkatachalam

Abstract

Horseradish peroxidase (HRP), a plant glycoprotein with a molecular weight of 40,000 D and a molecular radius (ae) of 30 A, has been modified chemically to prepare tracer molecules with different molecular charge. Modification of free carboxyl groups on the enzyme is achieved by carbodiimide activation and subsequent reaction of activated carboxyl groups with a nucleophile; uncharged groups or radicals containing additional positively charged moieties are introduced into the protein molecule resulting in an increased net positive charge of the tracer. Amino groups in the protein molecule are modified by acetylation or succinylation; this reaction will increase the net negative charge of the enzyme by either introducing an uncharged group or an additional carboxyl radical. The tracer molecules so obtained are then characterized in terms of molecular size and charge by column chromatography and isoelectric focusing respectively. The enzymatic activity as measured by 3,3'-diaminobenzidine reaction, the pH optimum and the absorption spectra for the modified enzymes remain virtually unchanged.

References

Sep 5, 2003·Journal of Immunoassay & Immunochemistry·Tulsidas G Shrivastav
Feb 1, 1982·The Journal of Clinical Investigation·W A BorderA H Cohen
Feb 1, 1983·Experimental Cell Research·J NilssonJ Thyberg
Jul 2, 1998·Acta Physiologica Scandinavica·J SörenssonB Haraldsson
Feb 21, 2013·Journal of Materials Chemistry. B, Materials for Biology and Medicine·Yanxia LiGuonan Chen
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