Spin-label study of energy-coupled active transport in Escherichia coli membrane vesicles

Biochemistry ◽  
1974 ◽  
Vol 13 (25) ◽  
pp. 5210-5214 ◽  
Author(s):  
Joseph J. Baldassare ◽  
Dan E. Robertson ◽  
Alice G. McAfee ◽  
Chien Ho
Keyword(s):  
Escherichia Coli ◽  
Active Transport ◽  
Spin Label ◽  
Membrane Vesicles ◽  
Label Study

The Orientation of Cytochrome c Oxidase in Coupled Phospholipid Membrane Vesicles—A Spin-Label Study

1975 ◽  
Vol 53 (3) ◽  
pp. 364-370 ◽  
Author(s):  
J. A. Kornblatt ◽  
W. L. Chen ◽  
J. C. Hsia ◽  
G. R. Williams
Keyword(s):  
Molecular Weight ◽  
Binding Site ◽  
Cytochrome Oxidase ◽  
Spin Label ◽  
Resonance Spectrum ◽  
Membrane Vesicles ◽  
Phospholipid Membrane ◽  
Vesicle Membrane ◽  
Exterior Surface ◽  
Label Study

Cytochrome oxidase, an enzyme containing six different subunits, has been shown to span the inner mitochrondrial membrane. The arrangement of the subunits within the membrane is unknown. We have specifically labeled the 25 000 molecular weight subunit with a spin-label derivative of N-ethylmaleimide, 3-maleimido-2,2,5,5-tetramethyl-1-pyrrolidinyloxyl (NEM-SL(5)). NEM-SL(5)-labeled cytochrome oxidase can be incorporated into phospholipid membranes to form coupled vesicles of the Hinkle, Kim &Racker ((1972) J. Biol. Chem. 247, 1338–1339) type. The resonance spectrum of NEM-SL(5) is similar in both soluble and vesicular cytochrome oxidase. Since ascorbate has been shown to reduce only spin label that is exposed to the exterior surface of a closed vesicle, we have used ascorbate to determine the NEM-SL(5)-binding site in the coupled vesicles. NEM-SL(5)-labeled cytochrome oxidase vesicles are reduced by 10 mM ascorbate with [Formula: see text] of 1 min at 22 °C. The rate of reduction is relatively independent of temperature. We conclude that (1) cytochrome oxidase is unidirectionally or preferentially oriented in the vesicle membrane, and (2) the NEM-SL(5)-binding site on the 25 000 molecular weight subunit is exposed to the external aqueous medium.

A spin label study of E. coli membrane vesicles

1973 ◽  
Vol 53 (2) ◽  
pp. 617-623 ◽  
Author(s):  
Joseph J. Baldassare ◽  
Alice Gerke McAfee ◽  
Chien Ho
Keyword(s):  
Spin Label ◽  
Membrane Vesicles ◽  
Label Study ◽  
E Coli

Lateral diffusion, protein mobility, and phase transitions in Escherichia coli membranes. Spin label study

Biochemistry ◽  
1973 ◽  
Vol 12 (26) ◽  
pp. 5360-5369 ◽  
Author(s):  
Erich Sackmann ◽  
Hermann Traeuble ◽  
Hans J. Galla ◽  
Peter Overath
Keyword(s):  
Escherichia Coli ◽  
Phase Transitions ◽  
Spin Label ◽  
Lateral Diffusion ◽  
Protein Mobility ◽  
Label Study
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