To the top

Page Manager: Webmaster
Last update: 9/11/2012 3:13 PM

Tell a friend about this page
Print version

Properties of the apo-for… - University of Gothenburg, Sweden Till startsida
Sitemap
To content Read more about how we use cookies on gu.se

Properties of the apo-form of the NADP(H)-binding domain III of proton-pumping Escherichia coli transhydrogenase: implications for the reaction mechanism of the intact enzyme

Journal article
Authors Anders Pedersen
J Karlsson
Magnus Althage
Jan Rydström
Published in Biochim.Biophys. Acta (Bioenergetics)
Volume 1604
Pages 55-59
Publication year 2003
Published at Department of Chemistry
Pages 55-59
Language en
Links dx.doi.org/10.1016/S0005-2728(03)00...
Keywords Transhydrogenase; NAD; NADP; Proton pump; Membrane protein
Subject categories Chemical Sciences

Abstract

Proton-translocating nicotinamide nucleotide transhydrogenases contain an NAD(H)-binding domain (dI), an NADP(H)-binding domain (dIII) and a membrane domain (dII) with the proton channel. Separately expressed and isolated dIII contains tightly bound NADP(H), predominantly in the oxidized form, possibly representing a so-called “occluded” intermediary state of the reaction cycle of the intact enzyme. Despite a Kd in the micromolar to nanomolar range, this NADP(H) exchanges significantly with the bulk medium. Dissociated NADP+ is thus accessible to added enzymes, such as NADP-isocitrate dehydrogenase, and can be reduced to NADPH. In the present investigation, dissociated NADP(H) was digested with alkaline phosphatase, removing the 2′-phosphate and generating NAD(H). Surprisingly, in the presence of dI, the resulting NADP(H)-free dIII catalyzed a rapid reduction of 3-acetylpyridine-NAD+ by NADH, indicating that 3-acetylpyridine-NAD+ and/or NADH interacts unspecifically with the NADP(H)-binding site. The corresponding reaction in the intact enzyme is not associated with proton pumping. It is concluded that there is a 2′-phosphate-binding region in dIII that controls tight binding of NADP(H) to dIII, which is not a required for fast hydride transfer. It is likely that this region is the Lys424–Arg425–Ser426 sequence and loops D and E. Further, in the intact enzyme, it is proposed that the same region/loops may be involved in the regulation of NADP(H) binding by an electrochemical proton gradent.

Page Manager: Webmaster|Last update: 9/11/2012
Share:

The University of Gothenburg uses cookies to provide you with the best possible user experience. By continuing on this website, you approve of our use of cookies.  What are cookies?