Evidence for allosterism in ribulose-1,5-bisphosphate carboxylase/oxygenase from comfrey
Evidence has been obtained suggesting that ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) is an allosteric enzyme in the sense that it shows cooperative active site binding, cooperative interactions between the activation and active sites and significant binding of some metabolites at a second site. Investigation of the binding of a potent competitive inhibitor. 2-carboxymannitol-1,6-bisphosphate (CMBP) by /sup 31/P-NMR indicated essentially 1:1 binding with the active sites of comfrey RuBisCo. Among the interactions of competitive inhibitors, as measured by difference UV spectroscopy, the binding curves for ortho-phosphate and ribose-5-phosphate were better fitted by a Monod-Wyman-Changeux model than by an independent site model, whereas the binding of CMBP and 2-phosphoglycolate were not. Difference UV methods also were used to study activation by CO/sub 2/ which at pH 7.9 in 10 mM MgCl/sub 2/ showed positive cooperativity with k = 100 +/- 3 ..mu..M (based on pK/sub a/ = 6.4 for the CO/sub 2/-HCO/sub 3//sup -/ equilibrium) and L = 3.5 +/- 0.7. Addition of saturating amounts of CMBP and lowering the MgCl/sub 2/ to 2 mM still gave a sigmoidal curve but it was shifted to higher CO/sub 2/ concentrations (k = 124 +/- 2 ..mu..M and L = 31 +/- 3). In the absence of CMBP the same conditions gave k = 26 +/- 2 ..mu..M for L = 3.5. Conversely, k was 0.96 +/- 0.08 ..mu..M for CMBP in 0.5 mM MgCl/sub 2/ without added NaHCO/sub 3/ but was 21 +/- 0.06 ..mu..M in 10 MgCl/sub 2/ and 2 mM NaHCO/sub 3/, pH 7.3.
- Research Organization:
- Kansas State Univ., Manhattan
- OSTI ID:
- 5180002
- Report Number(s):
- CONF-8606151-
- Journal Information:
- Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Vol. 45:6; Conference: 76. annual meeting of the Federation of American Society for Experimental Biology, Washington, DC, USA, 8 Jun 1986
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
62 RADIOLOGY AND NUCLEAR MEDICINE
OXYGENASES
BIOCHEMICAL REACTION KINETICS
CARBON DIOXIDE
HERBS
NMR SPECTRA
NUCLEAR MAGNETIC RESONANCE
PHOSPHATES
PHOSPHORUS 31
ULTRAVIOLET SPECTRA
CARBON COMPOUNDS
CARBON OXIDES
CHALCOGENIDES
ENZYMES
ISOTOPES
KINETICS
LIGHT NUCLEI
MAGNETIC RESONANCE
NUCLEI
ODD-EVEN NUCLEI
OXIDES
OXIDOREDUCTASES
OXYGEN COMPOUNDS
PHOSPHORUS COMPOUNDS
PHOSPHORUS ISOTOPES
PLANTS
REACTION KINETICS
RESONANCE
SPECTRA
STABLE ISOTOPES
550201* - Biochemistry- Tracer Techniques
550600 - Medicine