Interactions of haemoglobin with the Neisseria meningitidis receptor HpuAB: the role of TonB and an intact proton motive force.
We have characterized the interaction of the Neisseria meningitidis TonB-dependent receptor HpuAB with haemoglobin (Hb). Protease accessibility assays indicated that HpuA and HpuB are surface exposed, HpuB interacts physically with HpuA, and TonB energization affects the conformation of HpuAB. Binding assays using [125I]-Hb revealed that the bipartite receptor has a single binding site for Hb (Kd 150 nM). Competitive binding assays using heterologous Hbs revealed that HpuAB Hb recognition was not species specific. The binding kinetics of Hb to HpuAB were dramatically altered in a TonB- mutant and in wild-type meningococci treated with the protonophore carbonylcyanide m-chlorophenylhydrazone (CCCP), indicating that TonB and an intact proton motive force are required for normal Hb binding and release from HpuAB. Our results support a model in which both HpuA and HpuB are required to form a receptor complex in the outer membrane with a single binding site, whose structure and ligand interactions are significantly affected by the TonB-mediated energy state of the receptor.
Rohde
K H
KH
Department of Microbiology and Immunology, University of Oklahoma Health Sciences Center, Oklahoma City 73104, USA. kyle-rohde@ouhsc.edu
Gillaspy
A F
AF
Hatfield
M D
MD
Lewis
L A
LA
Dyer
D W
DW
D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism*; D006454:Hemoglobins / Q000378:metabolism*; D007700:Kinetics; D008024:Ligands; D008565:Membrane Proteins / Q000378:metabolism*; D009345:Neisseria meningitidis / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D006454:Hemoglobins; C445303:HpuA protein, Neisseria; C105625:HpuB protein, Neisseria; D008024:Ligands; D008565:Membrane Proteins; D011956:Receptors, Cell Surface; C057616:tonB protein, Bacteria; D014357:Trypsin