Bovine heart mitochondrial transhydrogenase catalyzes an exchange reaction between NADH and NAD+.

Wu, Licia N.Y.; Earle, Steven R.; Fisher, Ronald R.

doi:10.1016/s0021-9258(19)68977-5

Cited by 35 publications

(3 citation statements)

References 27 publications

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“…The fact that the rates of the forward and reverse transhydrogenation are not very different as cat-alyzed by the protein fragments lacking the enzyme's proton translocation domain II may indicate that in the intact enzyme this domain inhibits NADH 3 AcPyADP transhydrogenation in the absence of a proton motive force. The last entry in Table I shows transhydrogenation from NADH to AcPyAD, an activity that the intact enzyme does not exhibit unless under special conditions in the presence of NADP(H) (Wu et al, 1981;Fisher and Earle, 1982;Hutton et al, 1994). However, we have observed that the E. coli transhydrogenase mutant ␤H91K also catalyzes NADH to AcPyAD transhydrogenation in the absence of added NADP(H).…”

Section: Fig 1 Schematic Representation Of the Tridomain Composition Of The Bovine And The R Rubrum Transhydrogenases And The Bidomain Comentioning

confidence: 75%

Proton-translocating Nicotinamide Nucleotide Transhydrogenase

Yamaguchi¹,

Hatefi²

1995

Journal of Biological Chemistry

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The nicotinamide nucleotide transhydrogenase of bovine mitochondria is a homodimer of monomer M(r) = 109,065. The monomer is composed of three domains, an NH2-terminal 430-residue-long hydrophilic domain I that binds NAD(H), a central 400-residue-long hydrophobic domain II that is largely membrane intercalated and carries the enzyme's proton channel, and a COOH-terminal 200-residue-long hydrophilic domain III that binds NADP(H). Domains I and III protrude into the mitochondrial matrix, where they presumably come together to form the enzyme's catalytic site. The two-subunit transhydrogenase of Escherichia coli and the three-subunit transhydrogenase of Rhodospirillum rubrum have each the same overall tridomain hydropathy profile as the bovine enzyme. Domain I of the R. rubrum enzyme (the alpha 1 subunit) is water soluble and easily removed from the chromatophore membranes. We have isolated domain I of the bovine transhydrogenase after controlled trypsinolysis of the purified enzyme and have expressed in E. coli and purified therefrom domain III of this enzyme. This paper shows that an active bidomain transhydrogenase lacking domain II can be reconstituted by the combination of purified bovine domains I plus III or R. rubrum domain I plus bovine domain III.

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Section: Fig 1 Schematic Representation Of the Tridomain Composition Of The Bovine And The R Rubrum Transhydrogenases And The Bidomain Comentioning

confidence: 75%

Proton-translocating Nicotinamide Nucleotide Transhydrogenase

Yamaguchi¹,

Hatefi²

1995

Journal of Biological Chemistry

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“…Cyclic Reactions Catalyzed by Mutants and Wild-Type Enzyme in the Presence or Absence of rrI. The cyclic reduction of AcPyAD + by NADH is catalyzed by the NADP(H)-bound enzyme and occurs without a net proton transport ( , ). To detect effects of the redox state of NADP(H), this reaction was performed in the presence of NADP + or NADPH.…”

Section: Resultsmentioning

confidence: 99%

Identification of a Region Involved in the Communication between the NADP(H) Binding Domain and the Membrane Domain in Proton Pumping E. coli Transhydrogenase

2001

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The two hydrophilic domains I and III of Escherichia coli transhydrogenase containing the binding sites for NAD(H) and NADP(H), respectively, are located on the cytosolic side of the membrane, whereas the hydrophobic domain II is composed of 13 transmembrane alpha-helices, and is responsible for proton transport. In the present investigation the segment betaC260-betaS266 connecting domain II and III was characterized primarily because of its assumed role in the bioenergetic coupling of the transhydrogenase reaction. Each residue of this segment was replaced by a cysteine in a cysteine-free background, and the mutated proteins analyzed. Except for betaS266C, binding studies of the fluorescent maleimide derivative MIANS to each cysteine in the betaC260-betaR266 region revealed an increased accessibility in the presence of NADP(H) bound to domain III; an opposite effect was observed for betaS266. A betaD213-betaR265 double cysteine mutant was isolated in a predominantly oxidized form, suggesting that the corresponding residues in the wild-type enzyme are closely located and form a salt bridge. The betaS260C, betaK261C, betaA262C, betaM263, and betaN264 mutants showed a pronounced inhibition of proton-coupled reactions. Likewise, several betaR265 mutants and the D213C mutant showed inhibited proton-coupled reactions but also markedly increased values. It is concluded that the mobile hinge region betaC260-betaS266 and the betaD213-betaR265 salt bridge play a crucial role in the communication between the proton translocation/binding events in domain II and binding/release of NADP(H) in domain III.

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“…It is a wholly artificial process: the reduction of AcPdAD + (an analogue of NAD + ) by NADH in the presence of NADP + /NADPH. It was discovered with mitochondrial transhydrogenase (73) and explained (74,75), though the reaction was thought erroneously to be coupled to proton translocation. Experiments with detergent-dispersed E. coli enzyme led to a refined description (59).…”

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confidence: 99%