Using 31P NMR and freeze‐fracture electron microscopy we investigated the effect of several synthetic signal peptides on lipid structure in model membranes mimicking the lipid composition of the Escherichia coli inner membrane. It is demonstrated that the signal peptide of the E. coli outer membrane protein PhoE, as well as that of the M13 phage coat protein, strongly promote the formation of non‐bilayer lipid structures. This effect appears to be correlated to in vivo translocation efficiency, since a less functional analogue of the PhoE signal peptide was found to be less active in destabilizing the bilayer. It is proposed that signal sequences can induce local changes in lipid structure that are involved in protein translocation across the membrane.

Isolation and characterization of three subcomplexes of the mitochondrial NADH: ubiquinone oxidoreductase (complex I). Finel, M.; Majander, A.S.; Tyynela, J.; de Jong, A.M.P.; Albracht, S.P.J.; Wilkstrom, M. General rightsIt is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s) and/or copyright holder(s), other than for strictly personal, individual use, unless the work is under an open content license (like Creative Commons). Disclaimer/Complaints regulationsIf you believe that digital publication of certain material infringes any of your rights or (privacy) interests, please let the Library know, stating your reasons. In case of a legitimate complaint, the Library will make the material inaccessible and/or remove it from the website. Please Ask the Library: http://uba.uva.nl/en/contact, or a letter to: Library of the University of Amsterdam, Secretariat, Singel 425, 1012 WP Amsterdam, The Netherlands. You will be contacted as soon as possible. Download date: 10 May 2018Eur. J. Biochem. 226, 237-242 (1994) Enzymically active subcomplexes were purified from bovine mitochondrial NADH :ubiquinone oxidoreductase (complex I) by sucrose-gradient centrifugation in the presence of detergents. These subcomplexes, named 12, IS, and US, catalyse ferricyanide and ubiquinone-1 (Q-1) reduction by NADH at similar rates to complex I, but do not catalyse the reduction of decylubiquinone. In addition, the Q-1 reductase activity of all the subcomplexes is insensitive to rotenone. Chemical and EPR analyses of the subcomplexes show that FMN and all the Fe-S clusters of complex I are present, but that the line shape of cluster 2 is modified. The smallest subcomplex, US, contains only approximately 13 subunits, as compared to approximately 22 in the previously described sub-

Until now ubisemiquinones associated with NADH: ubiquinone oxidoreductase (complex I) have been reported to occur in isolated enzyme and in tightly coupled submitochondrial particles. In this report it is shown that ubisemiquinones are always detectable during steady‐state electron transfer from NADH to ubiquinone, independent of the type of inner‐membrane preparation used. The EPR signal of the rotenone‐sensitive ubisemiquinones could be detected not only in coupled MgATP submitochondrial particles, but also in routine preparations of uncoupled submitochondrial particles and in mitochondria. The ubisemiquinone formation in coupled preparations was completely insensitive to uncouplers. The maximal radical concentration during steady‐state electron transfer from NADH to quinone was equal to that of iron‐sulphur cluster 2. Experiments with antimycin, myxothiazol and 2‐thenoyltrifluoroacetone demonstrated that about half of this radical was associated with complex I, giving a ubisemiquinone concentration of about 0.5 mol semiquinone/mol cluster 2. Uncoupled submitochondrial particles, prepared by extensive sonification, never showed radical signals within 100 ms after mixing with NADH. This was due to the reversible inactivation of the enzyme, caused by elevated temperatures during sonification. In preparations with deliberately heat‐inactivated complex I, no radical signals were detected within 200 ms after mixing with NADH; at 1 s, however, radical formation was maximal. Yet, depending on the procedure of reactivation of the complex, in preparations previously treated to inactivate them ubisemiquinone concentrations were always less than in untreated particles. When complex I was in the active state the ubisemiquinone signal was maximal within 40 ms. The results described in this report lead to the conclusion that ubisemiquinones form obligatory intermediates in the reaction of NADH dehydrogenase with ubiquinone.

The reproducibility of the ACS-NL was high. The ACS-NL institutional version score may be biased in favour of men.