Helicobacter pylori neutrophil-activating protein activates neutrophils by its C-terminal region even without dodecamer formation, which is a prerequisite for DNA protection--novel approaches against Helicobacter pylori inflammation.
Helicobacter pylori neutrophil-activating protein (HP-NAP) protects DNA from free radicals as a dodecamer through its ferroxidase activity without, however, directly binding to it. The retardation that was observed at pH 7.5 could be easily attributed to an iron effect, as it was revealed by experiments in the absence of HP-NAP. A total loss of ferroxidase activity, dodecamer formation and DNA protection in environments rich in free radicals was observed after replacement of His25, His37, Asp52 and Lys134, which are located within the ferroxidase site, with Ala. Molecular dynamics simulations revealed that dimer formation is highly unlikely following mutation of the above amino acids, as the Fe(2+) is no longer attracted with equal strength by both subunits. These findings probably indicate that iron plays an important role in the conformation of HP-NAP by initiating the formation of stable dimers that are indispensable for the ensuing dodecamer structure. Very surprisingly, neutrophil activation appeared to be stimulated by structural elements that are localized within the C-terminal region of both mutant HP-NAP and wild-type dodecamer HP-NAP. In particular, the dodecamer conformation does not seem to be necessary for activation, and helices H3 (Leu69-Leu75) and H4 (Lys89-Leu114) or the linking coils (His63-Thr68 and Thr76-Ser88) are probably critical in stimulating neutrophil activation.
Kottakis
Filippos
F
Laboratory of Biochemistry, School of Chemistry, Aristotle University of Thessaloniki, TK 54124, Thessaloniki, Greece.
Papadopoulos
Georgios
G
Pappa
Eleni V
EV
Cordopatis
Paul
P
Pentas
Stefanos
S
Choli-Papadopoulou
Theodora
T
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology*; D001483:Base Sequence; D001704:Biopolymers / Q000378:metabolism*; D002570:Ceruloplasmin / Q000378:metabolism; D004249:DNA Damage; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D016481:Helicobacter Infections / Q000517:prevention & control*; D016480:Helicobacter pylori / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D017665:Hydroxyl Radical / Q000378:metabolism; D007501:Iron / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D018375:Neutrophil Activation / Q000502:physiology*; D009504:Neutrophils / Q000378:metabolism*
D001426:Bacterial Proteins; D001704:Biopolymers; D017931:DNA Primers; C092412:neutrophil-activating protein A, Helicobacter pylori; D017665:Hydroxyl Radical; D007501:Iron; D002570:Ceruloplasmin