Temperature effects on the MgATP-induced electron transfer between the nitrogenase proteins from Azotobacter vinelandii.
The temperature dependence of the pre-steady-state MgATP-dependent electron transfer from the MoFe protein to the Fe protein of the nitrogenase from Azotobacter vinelandii has been investigated between 6 degrees C and 31 degrees C by stopped-flow spectrophotometry. Below 14 degrees C, the data are consistent with a model in which interaction of MgATP with nitrogenase is fast and irreversible, and is followed by reversible electron transfer. From the extent and from the rate of the absorbance change, the rate constants for electron transfer from Fe protein to MoFe protein and of the reverse reaction were calculated. The direct rate constant increases with temperature (6-14 degrees C) from about 1 s-1 to about 26 s-1. The rate constant for the reverse reaction was found to be approximately 4 s-1 and invariant with the reaction temperature. Analysis of the data obtained in the temperature range between 6 degrees C and 12 degrees C within the framework of the transition-state theory show that electron transfer from the Fe protein to the MoFe protein occurs via a highly disordered transition state with activation parameters delta H(0) ++ = 289 kJ.mol-1 and delta S(0) ++ = 792 J.K-1.mol-1. The Eyring plot of the stopped-flow data displays an inflection point around 14 degrees C. From the stopped-flow data obtained between 18 degrees and 27 degrees C the activation parameters delta H(0) ++ and delta S(0) ++ for the reduction of the MoFe protein by Fe protein are calculated to be 90 kJ.mol-1 and 99 J.K-1.mol-1 respectively. A second inflection point in the Eyring plot could exist around 28 degrees C.
Mensink
R E
RE
Department of Biochemistry, Agricultural University, Wageningen, The Netherlands.
Haaker
H
H
D000255:Adenosine Triphosphate / Q000378:metabolism*; D016948:Azotobacter vinelandii / Q000201:enzymology*; D004579:Electron Transport; D007501:Iron; D007700:Kinetics; D008982:Molybdenum; D009591:Nitrogenase / Q000378:metabolism*; D010084:Oxidation-Reduction; D013053:Spectrophotometry; D013696:Temperature; D013816:Thermodynamics
D008982:Molybdenum; D000255:Adenosine Triphosphate; D007501:Iron; D009591:Nitrogenase