Gallium protoporphyrin as an antimicrobial for non-typeable Haemophilus influenzae in COPD patients.

AIMS: Colonisation with non-typeable Haemophilus influenzae (NTHi) is common in COPD. Iron is required by bacteria for nutrition. Gallium is imported into bacteria using iron import proteins. Gallium cannot fulfill key metabolic functions, causing bactericidal effects. We tested the efficacy of gallium compounds as antimicrobials against NTHi in hemin rich conditions, and their ability to reduce NTHi induced pro-inflammatory responses in macrophages. MAIN METHODS: NTHi was cultured with the free iron analogue gallium nitrate (GaN) and heme iron analogue gallium protoporphyrin (GaPP) (0.5-4 µM; 24 h). Growth of NTHi reference strain (NCTC 12699) and 6 clinical isolates from COPD patients (including antibiotic resistant isolates) was assessed by optical density, and viability by Miles Misra. Monocyte derived macrophages (MDMs) were treated with GaPP before/after NTHi exposure. Viable intracellular NTHi was assessed by gentamicin protection assay. GaN or GaPP was added to NTHi cultures prior to culture with MDMs. Cytokine gene expression (qPCR) and protein secretion (ELISA) were measured. KEY FINDINGS: NTHi growth and viability were reduced by GaPP but not GaN. GaPP inhibited growth of COPD isolates (4 µM: 87 % reduction). GaPP reduced intracellular viability of NTHi in macrophage infection models. MDM cytokine gene expression and protein secretion (TNF-α, IL-6 and CXCL8) in response to NTHi was reduced (82, 66 and 86 % for gene expression) when cultured with GaPP 4 µM. SIGNIFICANCE: GaPP is an effective antimicrobial for NTHi while GaN showed no effect on growth or viability. Culture of NTHi with GaPP also reduced the pro-inflammatory cytokine response in MDMs.

Overproduction, purification and preliminary X-ray diffraction analysis of YncE, an iron-regulated Sec-dependent periplasmic protein from Escherichia coli.

The yncE gene of Escherichia coli encodes a predicted periplasmic protein of unknown function. The gene is de-repressed under iron restriction through the action of the global iron regulator Fur. This suggests a role in iron acquisition, which is supported by the presence of the adjacent yncD gene encoding a potential TonB-dependent outer-membrane transporter. Here, the preliminary crystallographic structure of YncE is reported, revealing that it consists of a seven-bladed beta-propeller which resembles the corresponding domain of the ;surface-layer protein' of Methanosarcina mazei. A full structure determination is under way in order to provide insight into the function of this protein.