Towards Bacterial Resistance via the Membrane Strategy: Enzymatic, Biophysical and Biomimetic Studies of the Lipid cis-trans Isomerase of Pseudomonas aeruginosa.

The lipid cis-trans isomerase (Cti) is a periplasmic heme-c enzyme found in several bacteria including Pseudomonas aeruginosa, a pathogen known for causing nosocomial infections. This metalloenzyme catalyzes the cis-trans isomerization of unsaturated fatty acids in order to rapidly modulate membrane fluidity in response to stresses that impede bacterial growth. As a consequence, breakthrough in the elucidation of the mechanism of this metalloenzyme might lead to new strategies to combat bacterial antibiotic resistance.

Mutating the environment of heme b of E. coli cytochrome bd-I oxidase shifts its redox potential by 200 mV without inactivating the enzyme.

Cytochrome bd-I catalyzes the reduction of oxygen to water with the aid of hemes b, b and d. Here, effects of a mutation of E445, a ligand of heme b and of R448, hydrogen bonded to E445 are studied electrochemically in the E. coli enzyme.

pH-dependent kinetics of NO release from E. coli bd-I and bd-II oxidase reveals involvement of Asp/Glu58.

Escherichia coli contains two cytochrome bd oxidases, bd-I and bd-II. The structure of both enzymes is highly similar, but they exhibit subtle differences such as the accessibility of the active site through a putative proton channel. Here, we demonstrate that the duroquinol:dioxygen oxidoreductase activity of bd-I increased with alkaline pH, whereas bd-II showed a broad activity maximum around pH 7.

E. coli cytochrome bd-I requires Asp58 in the CydB subunit for catalytic activity.

The reduction of oxygen to water is crucial to life and a central metabolic process. To fulfil this task, prokaryotes use among other enzymes cytochrome bd oxidases (Cyt bds) that also play an important role in bacterial virulence and antibiotic resistance. To fight microbial infections by pathogens, an in-depth understanding of the enzyme mechanism is required.

Structure of Escherichia coli cytochrome bd-II type oxidase with bound aurachin D.

Cytochrome bd quinol:O oxidoreductases are respiratory terminal oxidases so far only identified in prokaryotes, including several pathogenic bacteria. Escherichia coli contains two bd oxidases of which only the bd-I type is structurally characterized. Here, we report the structure of the Escherichia coli cytochrome bd-II type oxidase with the bound inhibitor aurachin D as obtained by electron cryo-microscopy at 3 Å resolution.

Stabilization of the Highly Hydrophobic Membrane Protein, Cytochrome Oxidase, on Metallic Surfaces for Direct Electrochemical Studies.

The cytochrome oxidase catalyzes the reduction of oxygen to water in bacteria and it is thus an interesting target for electrocatalytic studies and biosensor applications. The oxidase is completely embedded in the phospholipid membrane. In this study, the variation of the surface charge of thiol-modified gold nanoparticles, the length of the thiols and the other crucial parameters including optimal phospholipid content and type, have been performed, giving insight into the role of these factors for the optimal interaction and direct electron transfer of an integral membrane protein.