Details

Autor(en) / Beteiligte

• Levine, Rodney L.
• Garland, Donita
• Oliver, Cynthia N.
• Amici, Adolfo
• Climent, Isabel
• Lenz, Anke-G.
• Ahn, Bong-Whan
• Shaltiel, Shmuel
• Stadtman, Earl R.

Titel

[49] Determination of carbonyl content in oxidatively modified proteins

Ist Teil von

• Methods in Enzymology, 1990, Vol.186, p.464-478

Ort / Verlag

San Diego, CA: Elsevier Science & Technology

Erscheinungsjahr

1990

Quelle

MEDLINE

Beschreibungen/Notizen

• This chapter discusses methods to determine carbonyl content in oxidatively modified proteins. The methods described are (1) reduction of the carbonyl group to an alcohol with tritiated borohydride; (2) reaction of the carbonyl group with 2,4-dinitrophenylhydrazine to form the 2,4-dinitrophenylhydrazone; (3) reaction of the carbonyl with fluorescein thiosemicarbazide to form the thiosemicarbazone; and (4) reaction of the carbonyl group with fluorescein amine to form a Schiff base followed by reduction to the secondary amine with cyanoborohydride. Van Poelje and Snell have also quantitated protein-bound pyruvoyl groups through formation of a Schiff base with p-aminobenzoic acid followed by reduction with cyanoborohydride. Although a systematic investigation has not appeared, this method should also be useful in detecting other protein-bound carbonyl groups. Carbonyl content of proteins is expressed as moles carbonyl/mole subunit for purified proteins of known molecular weight. For extracts, the results may be given as nanomoles carbonyl/milligram protein. For a protein having a molecular weight of 50,000, a carbonyl content of 1 mol carbonyl/mol protein corresponds to 20 nmol carbonyl/mg proteins.