PDBsum entry 1m8h

Oxidoreductase

PDB id: 1m8h

Contents

Protein chains

413 a.a. *

Ligands

SO4

HEM ×2

H4B ×2

6NI ×2

Waters ×89

* Residue conservation analysis

PDB id:

1m8h

Name:

Oxidoreductase

Title:

Inducible nitric oxide synthase with 6-nitroindazole bound

Structure:

Inducible nitric oxide synthase. Chain: a, b. Fragment: oxygenase domain. Synonym: nos, type ii, inducible nos, inos, macrophage nos, mac-nos. Engineered: yes

Source:

Mus musculus. House mouse. Organism_taxid: 10090. Expressed in: escherichia coli. Expression_system_taxid: 562.

Biol. unit:

Dimer (from PDB file)

Resolution:

2.85Å R-factor: 0.243 R-free: 0.294

Authors:

R.J.Rosenfeld,E.D.Garcin,K.Panda,G.Andersson,A.Aberg,A.V.Wallace, D.J.Stuehr,J.A.Tainer,E.D.Getzoff

Key ref:

R.J.Rosenfeld et al. (2002). Conformational changes in nitric oxide synthases induced by chlorzoxazone and nitroindazoles: crystallographic and computational analyses of inhibitor potency. Biochemistry, 41, 13915-13925. PubMed id: 12437348 DOI: 10.1021/bi026313j

Date:

24-Jul-02 Release date: 07-Aug-02

Protein chains

P29477  (NOS2_MOUSE) -  Nitric oxide synthase, inducible from Mus musculus

Seq: 1144 a.a.

Struc: 413 a.a.

Enzyme reactions

• Enzyme class: E.C.1.14.13.39 - nitric-oxide synthase (NADPH).
• Reaction: 2 L-arginine + 3 NADPH + 4 O2 + H⁺ = 2 L-citrulline + 2 nitric oxide + 3 NADP⁺ + 4 H2O

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

reference

DOI no: 10.1021/bi026313j

Biochemistry 41:13915-13925 (2002)

PubMed id: 12437348

Conformational changes in nitric oxide synthases induced by chlorzoxazone and nitroindazoles: crystallographic and computational analyses of inhibitor potency.

R.J.Rosenfeld, E.D.Garcin, K.Panda, G.Andersson, A.Aberg, A.V.Wallace, G.M.Morris, A.J.Olson, D.J.Stuehr, J.A.Tainer, E.D.Getzoff.

ABSTRACT

Nitric oxide is a key signaling molecule in many biological processes, making regulation of nitric oxide levels highly desirable for human medicine and for advancing our understanding of basic physiology. Designing inhibitors to specifically target one of the three nitric oxide synthase (NOS) isozymes that form nitric oxide from the L-Arg substrate poses a significant challenge due to the overwhelmingly conserved active sites. We report here 10 new X-ray crystallographic structures of inducible and endothelial NOS oxygenase domains cocrystallized with chlorzoxazone and four nitroindazoles: 5-nitroindazole, 6-nitroindazole, 7-nitroindazole, and 3-bromo-7-nitroindazole. Each of these bicyclic aromatic inhibitors has only one hydrogen bond donor and therefore cannot form the bidentate hydrogen bonds that the L-Arg substrate makes with Glu371. Instead, all of these inhibitors induce a conformational change in Glu371, creating an active site with altered molecular recognition properties. The cost of this conformational change is approximately 1-2 kcal, based on our measured constants for inhibitor binding to the wild-type and E371A mutant proteins. These inhibitors derive affinity by pi-stacking above the heme and replacing both intramolecular (Glu371-Met368) and intermolecular (substrate-Trp366) hydrogen bonds to the beta-sheet architecture underlying the active site. When bound to NOS, high-affinity inhibitors in this class are planar, whereas weaker inhibitors are nonplanar. Isozyme differences were observed in the pterin cofactor site, the heme propionate, and inhibitor positions. Computational docking predictions match the crystallographic results, including the Glu371 conformational change and inhibitor-binding orientations, and support a combined crystallographic and computational approach to isozyme-specific NOS inhibitor analysis and design.