Hemin uptake in Porphyromonas gingivalis: Omp26 is a hemin-binding surface protein.
A 26-kDa outer membrane protein (Omp26) has been proposed to play a role in hemin acquisition by Porphyromonas gingivalis (T. E. Bramanti and S. C. Holt, J. Bacteriol. 174:5827-5839, 1992). We studied [55Fe]hemin uptake in P. gingivalis grown under conditions of hemin starvation (Omp26 expressed on the outer membrane surface) and hemin excess (Omp26 not expressed on surface). [55Fe]hemin uptake occurred rapidly in hemin-starved cells which incorporated up to 70% of total [55Fe]hemin within 3 min. P. gingivalis grown under hemin-starved conditions or treated with the iron chelator 2,2'-bipyridyl to induce an iron stress took up six times more [55Fe]hemin than hemin-excess-grown cells. Polyclonal monospecific anti-Omp26 antibody added to hemin-starved cells inhibited [55Fe]hemin uptake by more than 50%, whereas preimmune serum had no effect. [55Fe]hemin uptake in hemin-starved P. gingivalis was inhibited (36 to 67%) in the presence of equimolar amounts of unlabeled hemin, protoporphyrin IX, zinz protoporphyrin, and Congo red dye but was not inhibited in the presence of non-hemin-containing iron sources. Heat shock treatment (45 degrees C) of hemin-excess-grown P. gingivalis (which cases translocation of Omp26 to the surface) increased [55Fe]hemin uptake by threefold after 3 min in comparison with cells grown at 37 degrees C. However, no [55Fe] hemin uptake beyond 3 min was observed in either hemin-excess-grown or hemin-starved cells exposed to heat shock. In experiments using heterobifunctional cross-linker analysis, hemin and selected porphyrins were cross-linked to Omp26 in hemin-starved P. gingivalis, but no cross-linking was seen with hemin-excess-grown cells. However, cross-linking of hemin to Omp26 was observed after heat shock treatment of hemin-excess-grown cells. Finally, anti-Omp26 antibody inhibited cross-linked of hemin to Omp26. These findings indicate that hemin binding and transport into P.gingivalis cell mediated by Omp26.
Bramanti
T E
TE
Department of Periodontics, University of Texas Health Science Center at San Antonio 78284-7894.
Holt
S C
SC
D001425:Bacterial Outer Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism*; D001692:Biological Transport / Q000187:drug effects; D002462:Cell Membrane / Q000378:metabolism; D003224:Congo Red / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006427:Hemin / Q000378:metabolism*; D006454:Hemoglobins / Q000494:pharmacology; D007501:Iron / Q000378:metabolism; D007504:Iron Radioisotopes; D007700:Kinetics; D008565:Membrane Proteins / Q000378:metabolism*; D008970:Molecular Weight; D016966:Porphyromonas gingivalis / Q000187:drug effects / Q000378:metabolism*; D011480:Protease Inhibitors / Q000494:pharmacology; D011524:Protoporphyrins / Q000494:pharmacology
D001425:Bacterial Outer Membrane Proteins; D006454:Hemoglobins; D007504:Iron Radioisotopes; D008565:Membrane Proteins; D011480:Protease Inhibitors; D011524:Protoporphyrins; C017803:zinc protoporphyrin; D003224:Congo Red; D006427:Hemin; C028025:protoporphyrin IX; D007501:Iron