Electron-Spin-Resonance Studies of the Lipoamide ‘Swinging Arm’ of the Pyruvate Dehydrogenase Multienzyme Complex of Escherichia coli

1978 ◽  
Vol 6 (1) ◽  
pp. 225-226 ◽  
Author(s):  
MARY C. AMBROSE GRIFFIN ◽  
WILLIAM G. GRIFFIN ◽  
RICHARD N. PERHAM
Keyword(s):  
Escherichia Coli ◽  
Electron Spin Resonance ◽  
Electron Spin ◽  
Pyruvate Dehydrogenase ◽  
Multienzyme Complex ◽  
Spin Resonance

scholarly journals The function of ubiquinone in Escherichia coli

1970 ◽  
Vol 117 (3) ◽  
pp. 551-562 ◽  
Author(s):  
G. B. Cox ◽  
N. A. Newton ◽  
F. Gibson ◽  
A. M. Snoswell ◽  
J. A. Hamilton
Keyword(s):  
Escherichia Coli ◽  
Electron Spin Resonance ◽  
Steady State ◽  
Electron Transport ◽  
Electron Spin ◽  
Electron Transport Chain ◽  
Electron Spin Resonance Signal ◽  
Resonance Signal ◽  
Transport Chain ◽  
Spin Resonance

1. The function of ubiquinone in Escherichia coli was studied by using whole cells and membrane preparations of normal E. coli and of a mutant lacking ubiquinone. 2. The mutant lacking ubiquinone, strain AN59 (Ubi−), when grown under aerobic conditions, gave an anaerobic type of growth yield and produced large quantities of lactic acid, indicating that ubiquinone plays a vital role in electron transport. 3. NADH and lactate oxidase activities in membranes from strain AN59 (Ubi−) were greatly impaired and activity was restored by the addition of ubiquinone (Q-1). 4. Comparison of the percentage reduction of flavin, cytochrome b1 and cytochrome a2 in the aerobic steady state in membranes from the normal strain (AN62) and strain AN59 (Ubi−) and the effect of respiratory inhibitors on these percentages in membranes from strain AN62 suggest that ubiquinone functions at more than one site in the electron-transport chain. 5. Membranes from strain AN62, in the absence of substrate, showed an electron-spin-resonance signal attributed to ubisemiquinone. The amount of reduced ubiquinone (50%) found after rapid solvent extraction is consistent with the existence of ubiquinone in membranes as a stabilized ubisemiquinone. 6. The effects of piericidin A on membranes from strain AN62 suggest that this inhibitor acts at the ubiquinone sites: thus inhibition of electron transport is reversed by ubiquinone (Q-1); the aerobic steady-state oxidation–reduction levels of flavins and cytochrome b1 in the presence of the inhibitor are raised to values approximating those found in the membranes of strain AN59 (Ubi−); the inhibitor rapidly eliminates the electron-spin-resonance signal attributed to ubisemiquinone and allows slow oxidation of endogenous ubiquinol in the absence of substrate and prevents reduction of ubiquinone in the presence of substrate. It is concluded that piericidin A separates ubiquinone from the remainder of the electron-transport chain. 7. A scheme is proposed in which ubisemiquinone, complexed to an electron carrier, functions in at least two positions in the electron-transport sequence.

scholarly journals Spin-label study of the mobility of enzyme-bound lipoic acid in the pyruvate dehydrogenase multienzyme complex of Escherichia coli

1976 ◽  
Vol 155 (2) ◽  
pp. 429-432 ◽  
Author(s):  
M C. Ambrose ◽  
R N. Perham
Keyword(s):  
Escherichia Coli ◽  
Lipoic Acid ◽  
Pyruvate Dehydrogenase ◽  
Active Sites ◽  
Catalytic Mechanism ◽  
Resonance Spectrum ◽  
Protein Molecule ◽  
Multienzyme Complex ◽  
Label Study ◽  
Spin Resonance

The lipoic acid residues covalently bound to the transacetylase component of the pyruvate dehydrogenase multienzyme complex of Escherichia coli were selectively modified by reaction with 4-maleimido-2,2,6,6-tetramethylpiperidino-oxyl. The electron-spin-resonance spectrum of the spin-labelled enzyme indicates that the bound nitroxide groups have high mobilities relative to the protein molecule. This physicochemical evidence is consistent with the view that the dithiolane ring of a lipoyl residue is capable of rapid migration between the active sites of the component enzymes in the catalytic mechanism.

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