Electron transport to nitrogenase in Azotobacter vinelandii

The enzyme nitrogenase requires MgATP, an anaerobic environment and an electron donor with a low redox potential for activity. The experiments described in this thesis deal with the electron transport to nitrogenase in Azotobacter vinelandii . It has been shown previously that the flow of reducing equivalents to nitrogenase is regulated by the Δψcomponent of the proton motive force. Short-term inhibition of nitrogenase activity by externally added NH4 +would be caused by lowering the Δψ. In Chapter 2 it has been shown that the extent of inhibition by NH 4 Cl is variable and depends upon the incubation conditions of the cells. Conditions are described, where nitrogenase activity is hardly inhibited by addition of NH 4 Cl and also conditions, where uptake of NH 4 Cl results in complete inhibition of nitrogenase activity. These results are discussed in Chapter 6. In addition to the membrane potential glutamine is proposed as another regulator of electron transport to nitrogenase.In Chapter 3 it has been shown, that whole cell nitrogenase activity is determined by the generation of reducing equivalents for nitrogenase. The physiological electron transport system to nitrogenase is very effective compared to the electron donor dithionite often used in invitro experiments. It has been shown that whole cell nitrogenase activity invivo can be twice the activity measured invitro . The consequences of this finding are discussed with respect to the mechanism for nitrogenase catalysis invitro and invivo .In Chapter 4 it has been shown, that three different flavodoxins can be isolated from A.vinelandii cells. Experimental evidence indicates that only flavodoxin II is involved in N 2 -fixation. The concentration of flavodoxin II is tenfold higher in N 2 -fixing cells compared to cells grown on NH 4 Ac. And its synthesis seems to be under the same regulatory control as the nitrogenase proteins.In Chapter 5 evidence is presented that a membrane bound NADPH dehydrogenase and two membrane bound polypeptides of relative molecular mass 29000 and 30000 probably play a role in electron transport to nitrogenase. Furthermore In Chapter 5 it has been demonstrated that there is a linear relationship between nitrogenase activity and the rate of respiration of A.vinelandii cells. It is proposed that the generation of reducing equivalents for nitrogenase is directly controlled by electron transfer activity in the respiratory chain.In Chapter 6 the new findings on the electron transport to nitrogenase are summarized in a scheme. In the scheme the electron carrier flavodoxin II is reduced by a membrane bound NADPH dehydrogenase only when the respiratory chain is functioning.

Bibliographic Details

Main Author:	Klugkist, J.
Other Authors:	Veeger, C.
Format:	Doctoral thesis biblioteca
Language:	English
Published:	Landbouwhogeschool
Subjects:	assimilation, azotobacter, biochemistry, metabolism, microorganisms, nitrogen, nitrogenase, synthesis, assimilatie, biochemie, metabolisme, micro-organismen, stikstof, synthese,
Online Access:	https://research.wur.nl/en/publications/electron-transport-to-nitrogenase-in-azotobacter-vinelandii
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