Bacterial outer membrane

Enzyme

In silico

Iron-binding proteins

Membrane

Escherichia coli

Biology

Gene

Membrane protein

Protein family

Molecular biology

Chemistry

Heme

Helicobacter pylori

Lactoferrin

Recombinant DNA

Genetics

Biochemistry

Introduction: Helicobacter pylori is a gram-negative spiral bacterial, has been associated with peptic ulcers, gastritis, duodenitis, and is believed to be the causative agent of gastric cancer. The sources such as human lactoferrin, haem and haemoglobin can support the H. pylori growth. However, still not fully understood how the process of iron acquisition occurs. An in silico analysis has shown that H. pylori genome has a family of three outer membrane protein regulated by iron (FrpB). Two of them: FrpB1 and FrpB2 were purified as recombinant proteins and their haem- or haemoglobin-binding capability was demonstrated. Unfortunately the last protein of the family (FrpB3) has not been investigated. Methods: In this work FrpB3 was purified by haem-affinity chromatography and its capacity of haem-binding was analyzed. This protein was identified by mass spectrometry and its expression was quantified by real time technique under different human iron sources. This expression was compared with frpB1 and frpb2. The FrpB3 structure was analyzed by 3D model to view the motifs necessary for Hb-binding, and also was compared with FrpB1 and FrpB2 structures. Results: The protein identified was FrpB3, its respective gene was overexpressed with haemoglobin. FrpB1 was overexpressed with haem while FrpB2 was induced in presence of haem and also haemoglobin. Both 3D models showed that they are structurally conserved because they have the typical barrel structure, which is inserted in membrane, also, the motifs necessary for Hb-binding were identified in all the structures. Conclusion: H. pylori express FrpB1, FrpB2 and FrpB3 proteins to scavenge iron and they are regulated according to availability of iron source, maybe in order to withstand the extreme environment present in the stomach. Our overall results represent the effort to explain the importance of iron acquisition.

Identification and Characterization of a Family of Outer Membrane Proteins of Helicobacter pylori, which Scavenge Iron from Human Sources