PDBsum entry 2frd

Oxidoreductase

PDB id: 2frd

Contents

Protein chains

360 a.a. *

174 a.a. *

Ligands

NAI ×2

NDP

Waters ×2

* Residue conservation analysis

PDB id:

2frd

Name:

Oxidoreductase

Title:

Structure of transhydrogenase (di.S138a.Nadh)2(diii.NADPH)1 asymmetric complex

Structure:

NAD(p) transhydrogenase subunit alpha part 1. Chain: a, b. Synonym: pyridine nucleotide transhydrogenase subunit alpha 1, nicotinamide nucleotide transhydrogenase subunit alpha 1, proton- translocating transhydrogenase component 1, di. Engineered: yes. Mutation: yes. NAD(p) transhydrogenase subunit beta. Chain: c.

Source:

Rhodospirillum rubrum. Organism_taxid: 1085. Gene: pntaa, nnta1. Expressed in: escherichia coli. Expression_system_taxid: 562. Gene: pntb, nntb. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008.

Biol. unit:

Trimer (from PQS)

Resolution:

3.20Å R-factor: 0.219 R-free: 0.272

Authors:

T.H.Brondijk,G.I.Van Boxel,O.C.Mather,P.G.Quirk,S.A.White,J.B.Jackson

Key ref:

T.H.Brondijk et al. (2006). The role of invariant amino acid residues at the hydride transfer site of proton-translocating transhydrogenase. J Biol Chem, 281, 13345-13354. PubMed id: 16533815 DOI: 10.1074/jbc.M513230200

Date:

19-Jan-06 Release date: 28-Feb-06

Protein chains

Q2RSB2  (PNTAA_RHORT) -  NAD(P) transhydrogenase subunit alpha part 1 from Rhodospirillum rubrum (strain ATCC 11170 / ATH 1.1.1 / DSM 467 / LMG 4362 / NCIMB 8255 / S1)

Seq: 384 a.a.

Struc: 360 a.a.*

Protein chain

P0C188  (PNTB_RHORU) -  NAD(P) transhydrogenase subunit beta from Rhodospirillum rubrum

Seq: 464 a.a.

Struc: 174 a.a.

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: Chains A, B, C: E.C.7.1.1.1 - proton-translocating NAD(P)(+) transhydrogenase.
• Reaction: NAD⁺ + NADPH + H⁺(in) = NADH + NADP⁺ + H⁺(out)

NAD(+) + NADPH (Bound ligand (Het Group name = NAI) corresponds exactly) + H(+)(in) (Bound ligand (Het Group name = NDP) corresponds exactly) = NADH + NADP(+) + H(+)(out)

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1074/jbc.M513230200

J Biol Chem 281:13345-13354 (2006)

PubMed id: 16533815

The role of invariant amino acid residues at the hydride transfer site of proton-translocating transhydrogenase.

T.H.Brondijk, G.I.van Boxel, O.C.Mather, P.G.Quirk, S.A.White, J.B.Jackson.

ABSTRACT

Transhydrogenase couples proton translocation across a membrane to hydride transfer between NADH and NADP+. Previous x-ray structures of complexes of the nucleotide-binding components of transhydrogenase ("dI2dIII1" complexes) indicate that the dihydronicotinamide ring of NADH can move from a distal position relative to the nicotinamide ring of NADP+ to a proximal position. The movement might be responsible for gating hydride transfer during proton translocation. We have mutated three invariant amino acids, Arg-127, Asp-135, and Ser-138, in the NAD(H)-binding site of Rhodospirillum rubrum transhydrogenase. In each mutant, turnover by the intact enzyme is strongly inhibited. Stopped-flow experiments using dI2dIII1 complexes show that inhibition results from a block in the steps associated with hydride transfer. Mutation of Asp-135 and Ser-138 had no effect on the binding affinity of either NAD+ or NADH, but mutation of Arg-127 led to much weaker binding of NADH and slightly weaker binding of NAD+. X-ray structures of dI2dIII1 complexes carrying the mutations showed that their effects were restricted to the locality of the bound NAD(H). The results are consistent with the suggestion that in wild-type protein movement of the Arg-127 side chain, and its hydrogen bonding to Asp-135 and Ser-138, stabilizes the dihydronicotinamide of NADH in the proximal position for hydride transfer.

Selected figure(s)

Figure 1.
FIGURE 1. The component structure of transhydrogenase. The gray shaded regions represent the dI[2]dIII[1] complex. The two dI polypeptides each comprise two domains, dI.1 (residues 1-137 and 328-384 in R. rubrum) and dI.2 (residues 138-327), which are separated by a deep cleft. The dIII component comprises a single domain. The dI polypeptide in the complex whose cleft is not associated with dIII is designated dI(A), and the dI polypeptide which is closely associated with dIII is designated dI(B). The predicted organization of transhydrogenase components that are absent in the dI[2]dIII[1] complex is shown by the dotted lines. The dashed arrows depict "forward" transhydrogenation and the thick solid arrows, the corresponding direction of proton translocation.

Figure 6.
FIGURE 6. Detail of the hydride transfer site of wild-type dI[2]dIII[1] complex and (dI.S138A). Atoms are shown in standard colors except C-4(N) atoms of the nicotinamide rings which are shown in pale yellow. A, detail from the dI(A) polypeptide of wild-type dI[2]dIII[1] complex (PDB, 1U2D (20)). The dihydronicotinamide ring of the bound NADH is in the distal position. B, detail from the interface of the dI(B) polypeptide and the dIII polypeptide of 1U2D. The dihydronicotinamide ring of the bound NADH is in the proximal position relative to the NADPH in dIII. C, detail from the interface of the dI(B) polypeptide and the dIII polypeptide of (dI.S138A)[2]dIII[1] complex (2FRD). The 2F[o]-F[c] electron density is contoured at 0.75 . Density for the dihydronicotinamide ring of the bound NADH is weak but indicates a distal position, as shown by the nucleotide in standard atom colors (but yellow for C-4(N)). For comparison, the nucleotide in cyan is in a proximal position; it is taken from the dI(B) polypeptide of 1U2D and superimposed at the adenine ring. The views in A-C are selected such that -sheet positions of the dI.2 domains superimpose and are shown with the adenine rings of the NADH in an alignment (black dashed line). The green dotted lines represent H-bonds. The polypeptide chain of dIII in (B and C) is not shown.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2006, 281, 13345-13354) copyright 2006.

Figures were selected by an automated process.