PDBsum entry 2bc1

Oxidoreductase

PDB id: 2bc1

Contents

Protein chains

473 a.a. *

Ligands

FAD ×2

Waters ×385

* Residue conservation analysis

PDB id:

2bc1

Name:

Oxidoreductase

Title:

Structural analysis of streptococcus pyogenes nadh oxidase: c44s nox

Structure:

Nadh oxidase. Chain: a, b. Engineered: yes. Mutation: yes

Source:

Streptococcus pyogenes. Organism_taxid: 1314. Expressed in: escherichia coli. Expression_system_taxid: 562.

Biol. unit:

Dimer (from PQS)

Resolution:

2.15Å R-factor: 0.198 R-free: 0.241

Authors:

W.H.Boles,T.C.Mallett

Key ref:

J.R.Wallen et al. (2015). Structural Analysis of Streptococcus pyogenes NADH Oxidase: Conformational Dynamics Involved in Formation of the C(4a)-Peroxyflavin Intermediate. Biochemistry, 54, 6815-6829. PubMed id: 26506002 DOI: 10.1021/acs.biochem.5b00676

Date:

18-Oct-05 Release date: 31-Oct-06

Protein chains

Q5XC60  (NAOX_STRP6) -  Probable NADH oxidase from Streptococcus pyogenes serotype M6 (strain ATCC BAA-946 / MGAS10394)

Seq: 456 a.a.

Struc: 473 a.a.*

* PDB and UniProt seqs differ at 3 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.7.1.1.2 - NADH:ubiquinone reductase (H(+)-translocating).
• Reaction: a ubiquinone + NADH + 5 H⁺(in) = a ubiquinol + NAD⁺ + 4 H⁺(out)

ubiquinone + NADH + 5 × H(+)(in) (Bound ligand (Het Group name = FAD) matches with 76.36% similarity) = ubiquinol + NAD(+) + 4 × H(+)(out)

• Cofactor: FMN; Iron-sulfur

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

reference

DOI no: 10.1021/acs.biochem.5b00676

Biochemistry 54:6815-6829 (2015)

PubMed id: 26506002

Structural Analysis of Streptococcus pyogenes NADH Oxidase: Conformational Dynamics Involved in Formation of the C(4a)-Peroxyflavin Intermediate.

J.R.Wallen, T.C.Mallett, T.Okuno, D.Parsonage, H.Sakai, T.Tsukihara, A.Claiborne.

ABSTRACT

In probing the oxygen reactivity of an Enterococcus faecalis NADH oxidase (Nox; O2 → 2H2O) C42S mutant lacking the Cys42-sulfenic acid (Cys42-SOH) redox center, we provided direct evidence of a C(4a)-peroxyflavin intermediate in the oxidative half-reaction and also described a conformational or chemical change that is rate-limiting for full reoxidation of the homodimer. In this work, the Nox from Streptococcus pyogenes (SpyNox) has been expressed and crystallized, and the overoxidized wild-type [Cys44-SOH → Cys44-sulfinic acid (Cys44-SO2H)] and C44S mutant enzyme structures have been refined at 2.0 and 2.15 Å, respectively. We show that azide binds to the two-electron reduced wild-type (EH2) enzyme and to the mutant enzyme in solution, but with a significantly higher affinity for the mutant protein. The spectral course of the titration with the SpyNox EH2 form clearly indicates progressive displacement of the Cys44-S(-) → FAD charge-transfer interaction. An azide soak with C44S Nox crystals led to the structure of the complex, as refined at 2.10 Å. The active-site N3(-) ligand is proximal to the Ser44 and His11 side chains, and a significant shift in the Ser44 side chain also appears. This provides an attractive explanation for the azide-induced loss of charge-transfer absorbance seen with the wild-type EH2 form and also permits accommodation of a C(4a)-peroxyflavin structural model. The conformation of Ser44 and the associated helical element, and the resulting steric accommodation, appear to be linked to the conformational change described in the E. faecalis C42S Nox oxidative half-reaction.