PDBsum entry 5amm

Oxidoreductase

PDB id: 5amm

Contents

Protein chains

267 a.a.

Ligands

HEM ×2

Metals

_CA ×2

__K ×2

Waters ×95

PDB id:

5amm

Name:

Oxidoreductase

Title:

Structure of leishmania major peroxidase d211n mutant

Structure:

Ascorbate peroxidase. Chain: a, b. Fragment: c-terminal catalytic domain, residues 35-303. Synonym: cytochromE C peroxidase. Engineered: yes. Mutation: yes

Source:

Leishmania major. Organism_taxid: 5664. Strain: friedlin. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

2.09Å R-factor: 0.187 R-free: 0.245

Authors:

G.Chreifi,J.B.Fields,S.A.Hollingsworth,M.Heyden,A.P.Arce,H.I.Magana- Garcia,T.L.Poulos,D.J.Tobias

Key ref:

J.B.Fields et al. (2015). "Bind and Crawl" Association Mechanism of Leishmania major Peroxidase and Cytochrome c Revealed by Brownian and Molecular Dynamics Simulations. Biochemistry, 54, 7272-7282. PubMed id: 26598276 DOI: 10.1021/acs.biochem.5b00569

Date:

11-Mar-15 Release date: 09-Dec-15

Protein chains

Q4Q3K2  (Q4Q3K2_LEIMA) -  Ascorbate peroxidase from Leishmania major

Seq: 303 a.a.

Struc: 267 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.1.11.1.11 - L-ascorbate peroxidase.
• Reaction: L-ascorbate + H2O2 = L-dehydroascorbate + 2 H2O
• Cofactor: Heme

Heme (Bound ligand (Het Group name = HEM) matches with 95.45% similarity)

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

reference

DOI no: 10.1021/acs.biochem.5b00569

Biochemistry 54:7272-7282 (2015)

PubMed id: 26598276

"Bind and Crawl" Association Mechanism of Leishmania major Peroxidase and Cytochrome c Revealed by Brownian and Molecular Dynamics Simulations.

J.B.Fields, S.A.Hollingsworth, G.Chreifi, M.Heyden, A.P.Arce, H.I.Magaña-Garcia, T.L.Poulos, D.J.Tobias.

ABSTRACT

Leishmania major, the parasitic causative agent of leishmaniasis, produces a heme peroxidase (LmP), which catalyzes the peroxidation of mitochondrial cytochrome c (LmCytc) for protection from reactive oxygen species produced by the host. The association of LmP and LmCytc, which is known from kinetics measurements to be very fast (∼10(8) M(-1) s(-1)), does not involve major conformational changes and has been suggested to be dominated by electrostatic interactions. We used Brownian dynamics simulations to investigate the mechanism of formation of the LmP-LmCytc complex. Our simulations confirm the importance of electrostatic interactions involving the negatively charged D211 residue at the LmP active site, and reveal a previously unrecognized role in complex formation for negatively charged residues in helix A of LmP. The crystal structure of the D211N mutant of LmP reported herein is essentially identical to that of wild-type LmP, reinforcing the notion that it is the loss of charge at the active site, and not a change in structure, that reduces the association rate of the D211N variant of LmP. The Brownian dynamics simulations further show that complex formation occurs via a "bind and crawl" mechanism, in which LmCytc first docks to a location on helix A that is far from the active site, forming an initial encounter complex, and then moves along helix A to the active site. An atomistic molecular dynamics simulation confirms the helix A binding site, and steady state activity assays and stopped-flow kinetics measurements confirm the role of helix A charges in the association mechanism.