Exploring by Pulsed EPR the Electronic Structure of Ubisemiquinone Bound at the QH Site of Cytochrome bo3 from Escherichia coli with in Vivo 13C-Labeled Methyl and Methoxy Substituents | Sciact - CRIS-система Института катализа

Exploring by Pulsed EPR the Electronic Structure of Ubisemiquinone Bound at the QH Site of Cytochrome bo3 from Escherichia coli with in Vivo 13C-Labeled Methyl and Methoxy Substituents Научная публикация

Журнал

Journal of Biological Chemistry
ISSN: 0021-9258 , E-ISSN: 1083-351X

Вых. Данные

Год: 2011, Том: 286, Номер: 12, Страницы: 10105-10114 Страниц : 10 DOI: 10.1074/jbc.M110.206821

Ключевые слова

Amino acids; Catalyst activity; Cell membranes; Electronic structure; Escherichia coli; Functional groups; Proteins

Авторы

Lin Myat T. ¹ , Shubin Alexander A. ³ , Samoilova Rimma I. ⁴ , Narasimhulu Kuppala V. ² , Baldansuren Amgalanbaatar ¹ , Gennis Robert B. ¹ , Dikanov Sergei A. ²

Организации

1	Department of Biochemistry, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801
2	Department of Veterinary Clinical Medicine, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801
3	Boreskov Institute of Catalysis, RussianAcademy of Sciences, Novosibirsk 630090, Russia
4	Institute of Chemical Kinetics and Combustion, RussianAcademy of Sciences, Novosibirsk 630090, Russia

Информация о финансировании (4)

1	National Institutes of Health	GM 62954
2	National Institutes of Health	S10-RR15878
3	United States Department of Energy	DE-FG02-08ER15960
4	United States Department of Energy	DE-FG02-87ER13716

The cytochromebo3ubiquinol oxidase from Escherichia coli resides in the bacterial cytoplasmic membrane and catalyzes the two-electron oxidation of ubiquinol-8 and four-electron reduc-tion of O2 to water. The one-electron reduced semiquinone forms transiently during the reaction, and the enzyme has been demonstrated to stabilize the semiquinone. The semiquinone is also formed in the D75E mutant, where the mutation has little influence on the catalytic activity, and in the D75H mutant, which is virtually inactive. In this work, wild-type cytochrome bo3 as well as the D75E and D75H mutant proteins were pre-pared with ubiquinone-8 13C-labeled selectively at the methyl and two methoxy groups. This was accomplished by expressing the proteins in a methionine auxotroph in the presence ofL-me-thionine with the side chain methyl group13C-labeled. The13C-labeled quinone isolated from cytochromebo3was also used for the generation of model anion radicals in alcohol. Two-dimen-sionalpulsedEPRandENDORwereusedforthestudyofthe13C methyl and methoxy hyperfine couplings in the semiquinone generatedinthethreeproteinsindicatedaboveandinthemodel system. The data were used to characterize the transferred unpaired spin densities on the methyl and methoxy substituents and the conformations of the methoxy groups. In the wild type and D75E mutant, the constraints on the configurations of the methoxy side chains are similar, but the D75H mutant appears to have altered methoxy configurations, which could be related to the perturbed electron distribution in the semiquinone and the loss of enzymatic activity.

Lin M.T. , Shubin A.A. , Samoilova R.I. , Narasimhulu K.V. , Baldansuren A. , Gennis R.B. , Dikanov S.A.
Exploring by Pulsed EPR the Electronic Structure of Ubisemiquinone Bound at the QH Site of Cytochrome bo3 from Escherichia coli with in Vivo 13C-Labeled Methyl and Methoxy Substituents

Journal of Biological Chemistry. 2011. V.286. N12. P.10105-10114. DOI: 10.1074/jbc.M110.206821 WOS Scopus РИНЦ CAPlusCA OpenAlex

Полный текст от издателя

Поступила в редакцию:	25 нояб. 2010 г.
Принята к публикации:	13 янв. 2011 г.
Опубликована online:	19 янв. 2011 г.
Опубликована в печати:	25 мар. 2011 г.

Web of science:	WOS:000288547000025
Scopus:	2-s2.0-79953195121
РИНЦ:	16983874
Chemical Abstracts:	2011:348865
Chemical Abstracts (print):	154:479082
OpenAlex:	W1972996979

БД	Цитирований
Web of science	19
Scopus	21
РИНЦ	21
OpenAlex	20