Modification of histidine at the active site of spinach ribulose bisphosphate carboxylase

Ashok Saluja, Bruce A. McFadden

Research output: Contribution to journalArticle

28 Citations (Scopus)

Abstract

Ribulose 1,5-bisphosphate carboxylase from spinach was rapidly inactivated by diethylpyrocarbonate (DEP) at pH 7.0 and 30°C. The inactivation showed saturation kinetics with a half-inactivation time at saturating DEP equal to 0.1 minutes and KDEP = 7.4 mM. One substrate, ribulose bisphosphate, the product 3-phosphoglycerate and two competitive inhibitors protected against inactivation, thereby indicating that DEP modifies the active site. DEP-modified enzyme showed an increased absorption at 240 nm which was lost upon treatment with 0.4 M hydroxylamine. Most of the activity lost by DEP modification could be restored after treatment with 0.4 M hydroxylamine at 4°C. The results suggest that DEP modified 2 to 3 histidine residues per 70,000-dalton combination of large and small subunits. These residues are essential to catalysis by the carboxylase activity of ribulose bisphosphate carboxylase/oxygenase.

Original languageEnglish (US)
Pages (from-to)1091-1097
Number of pages7
JournalBiochemical and Biophysical Research Communications
Volume94
Issue number4
DOIs
StatePublished - Jun 30 1980
Externally publishedYes

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Diethyl Pyrocarbonate
Ribulose-Bisphosphate Carboxylase
Spinacia oleracea
Histidine
Catalytic Domain
Hydroxylamine
Oxygenases
Catalysis
Kinetics
Substrates
Enzymes

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Modification of histidine at the active site of spinach ribulose bisphosphate carboxylase. / Saluja, Ashok; McFadden, Bruce A.

In: Biochemical and Biophysical Research Communications, Vol. 94, No. 4, 30.06.1980, p. 1091-1097.

Research output: Contribution to journalArticle

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