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. 1995 Dec;63(12):4634–4641. doi: 10.1128/iai.63.12.4634-4641.1995

Binding and accumulation of hemin in Neisseria gonorrhoeae.

P J Desai 1, R Nzeribe 1, C A Genco 1
PMCID: PMC173666  PMID: 7591117

Abstract

The ability to utilize hemin and hemin-containing compounds for nutritional iron (Fe) uptake has been documented for several pathogenic bacteria. Neisseria gonorrhoeae can utilize free hemin as a source of Fe for growth; however, little is known concerning the mechanisms involved in hemin transport. In this study we have characterized the binding and accumulation of hemin by N. gonorrhoeae and defined the specificity of the gonococcal hemin receptor. N. gonorrhoeae F62 was grown in a chemically defined medium containing the iron chelator Desferal, and hemin transport was initiated by the addition of [59Fe]hemin (4.0 or 8.0 microM; specific activity, 7.0 Ci/mol). 59Fe uptake from radiolabeled hemin by N. gonorrhoeae was energy dependent, and 59Fe was shown to accumulate in the cell at a constant rate during logarithmic growth. However, we observed a decrease in the uptake of 59Fe from radiolabeled hemin when inorganic iron was present in the growth medium. Binding of 59Fe from radiolabeled hemin was inhibited by the addition of either cold hemin, hematoporphyrin, or hemoglobin, but not by ferric citrate. Although [14C]hemin was found to support the growth of N. gonorrhoeae, we did not detect the uptake of 14C from radiolabeled hemin. Extraction of the gonococcal periplasmic ferric binding protein (Fbp) from cultures grown with [59Fe]hemin indicated that a majority of the 59Fe was associated with the Fbp. Taken together, the results presented here indicate that hemin binds to a gonococcal outer membrane receptor through the protoporphyrin portion of the molecule and that following binding, iron is removed and transported into the cell, where it is associated with the gonococcal periplasmic ferric binding protein, Fbp.

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Selected References

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  1. Anderson J. E., Sparling P. F., Cornelissen C. N. Gonococcal transferrin-binding protein 2 facilitates but is not essential for transferrin utilization. J Bacteriol. 1994 Jun;176(11):3162–3170. doi: 10.1128/jb.176.11.3162-3170.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Angerer A., Gaisser S., Braun V. Nucleotide sequences of the sfuA, sfuB, and sfuC genes of Serratia marcescens suggest a periplasmic-binding-protein-dependent iron transport mechanism. J Bacteriol. 1990 Feb;172(2):572–578. doi: 10.1128/jb.172.2.572-578.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bagg A., Neilands J. B. Ferric uptake regulation protein acts as a repressor, employing iron (II) as a cofactor to bind the operator of an iron transport operon in Escherichia coli. Biochemistry. 1987 Aug 25;26(17):5471–5477. doi: 10.1021/bi00391a039. [DOI] [PubMed] [Google Scholar]
  4. Berish S. A., Chen C. Y., Mietzner T. A., Morse S. A. Expression of a functional neisserial fbp gene in Escherichia coli. Mol Microbiol. 1992 Sep;6(18):2607–2615. doi: 10.1111/j.1365-2958.1992.tb01438.x. [DOI] [PubMed] [Google Scholar]
  5. Berish S. A., Kapczynski D. R., Morse S. A. Nucleotide sequence of the Fbp gene from Neisseria meningitidis. Nucleic Acids Res. 1990 Aug 11;18(15):4596–4596. doi: 10.1093/nar/18.15.4596. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Berish S. A., Mietzner T. A., Mayer L. W., Genco C. A., Holloway B. P., Morse S. A. Molecular cloning and characterization of the structural gene for the major iron-regulated protein expressed by Neisseria gonorrhoeae. J Exp Med. 1990 May 1;171(5):1535–1546. doi: 10.1084/jem.171.5.1535. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Berish S. A., Subbarao S., Chen C. Y., Trees D. L., Morse S. A. Identification and cloning of a fur homolog from Neisseria gonorrhoeae. Infect Immun. 1993 Nov;61(11):4599–4606. doi: 10.1128/iai.61.11.4599-4606.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Beucher M., Sparling P. F. Cloning, sequencing, and characterization of the gene encoding FrpB, a major iron-regulated, outer membrane protein of Neisseria gonorrhoeae. J Bacteriol. 1995 Apr;177(8):2041–2049. doi: 10.1128/jb.177.8.2041-2049.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Biswas G. D., Sparling P. F. Characterization of lbpA, the structural gene for a lactoferrin receptor in Neisseria gonorrhoeae. Infect Immun. 1995 Aug;63(8):2958–2967. doi: 10.1128/iai.63.8.2958-2967.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Blanton K. J., Biswas G. D., Tsai J., Adams J., Dyer D. W., Davis S. M., Koch G. G., Sen P. K., Sparling P. F. Genetic evidence that Neisseria gonorrhoeae produces specific receptors for transferrin and lactoferrin. J Bacteriol. 1990 Sep;172(9):5225–5235. doi: 10.1128/jb.172.9.5225-5235.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Bramanti T. E., Holt S. C. Hemin uptake in Porphyromonas gingivalis: Omp26 is a hemin-binding surface protein. J Bacteriol. 1993 Nov;175(22):7413–7420. doi: 10.1128/jb.175.22.7413-7420.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Chen C. Y., Genco C. A., Rock J. P., Morse S. A. Physiology and metabolism of Neisseria gonorrhoeae and Neisseria meningitidis: implications for pathogenesis. Clin Microbiol Rev. 1989 Apr;2 (Suppl):S35–S40. doi: 10.1128/cmr.2.suppl.s35. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Cope L. D., Yogev R., Muller-Eberhard U., Hansen E. J. A gene cluster involved in the utilization of both free heme and heme:hemopexin by Haemophilus influenzae type b. J Bacteriol. 1995 May;177(10):2644–2653. doi: 10.1128/jb.177.10.2644-2653.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Cornelissen C. N., Biswas G. D., Tsai J., Paruchuri D. K., Thompson S. A., Sparling P. F. Gonococcal transferrin-binding protein 1 is required for transferrin utilization and is homologous to TonB-dependent outer membrane receptors. J Bacteriol. 1992 Sep;174(18):5788–5797. doi: 10.1128/jb.174.18.5788-5797.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Cornelissen C. N., Sparling P. F. Iron piracy: acquisition of transferrin-bound iron by bacterial pathogens. Mol Microbiol. 1994 Dec;14(5):843–850. doi: 10.1111/j.1365-2958.1994.tb01320.x. [DOI] [PubMed] [Google Scholar]
  16. Daskaleros P. A., Stoebner J. A., Payne S. M. Iron uptake in Plesiomonas shigelloides: cloning of the genes for the heme-iron uptake system. Infect Immun. 1991 Aug;59(8):2706–2711. doi: 10.1128/iai.59.8.2706-2711.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Dyer D. W., West E. P., Sparling P. F. Effects of serum carrier proteins on the growth of pathogenic neisseriae with heme-bound iron. Infect Immun. 1987 Sep;55(9):2171–2175. doi: 10.1128/iai.55.9.2171-2175.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Genco C. A., Berish S. A., Chen C. Y., Morse S., Trees D. L. Genetic diversity of the iron-binding protein (Fbp) gene of the pathogenic and commensal Neisseria. FEMS Microbiol Lett. 1994 Feb 15;116(2):123–129. doi: 10.1111/j.1574-6968.1994.tb06690.x. [DOI] [PubMed] [Google Scholar]
  19. Genco C. A., Chen C. Y., Arko R. J., Kapczynski D. R., Morse S. A. Isolation and characterization of a mutant of Neisseria gonorrhoeae that is defective in the uptake of iron from transferrin and haemoglobin and is avirulent in mouse subcutaneous chambers. J Gen Microbiol. 1991 Jun;137(6):1313–1321. doi: 10.1099/00221287-137-6-1313. [DOI] [PubMed] [Google Scholar]
  20. Genco C. A., Odusanya B. M., Brown G. Binding and accumulation of hemin in Porphyromonas gingivalis are induced by hemin. Infect Immun. 1994 Jul;62(7):2885–2892. doi: 10.1128/iai.62.7.2885-2892.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Genco C. A. Regulation of hemin and iron transport in Porphyromonas gingivalis. Adv Dent Res. 1995 Feb;9(1):41–47. doi: 10.1177/08959374950090010801. [DOI] [PubMed] [Google Scholar]
  22. Genco C. A., Simpson W., Forng R. Y., Egal M., Odusanya B. M. Characterization of a Tn4351-generated hemin uptake mutant of Porphyromonas gingivalis: evidence for the coordinate regulation of virulence factors by hemin. Infect Immun. 1995 Jul;63(7):2459–2466. doi: 10.1128/iai.63.7.2459-2466.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hanson M. S., Hansen E. J. Molecular cloning, partial purification, and characterization of a haemin-binding lipoprotein from Haemophilus influenzae type b. Mol Microbiol. 1991 Feb;5(2):267–278. doi: 10.1111/j.1365-2958.1991.tb02107.x. [DOI] [PubMed] [Google Scholar]
  24. Hanson M. S., Slaughter C., Hansen E. J. The hbpA gene of Haemophilus influenzae type b encodes a heme-binding lipoprotein conserved among heme-dependent Haemophilus species. Infect Immun. 1992 Jun;60(6):2257–2266. doi: 10.1128/iai.60.6.2257-2266.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Henderson D. P., Payne S. M. Characterization of the Vibrio cholerae outer membrane heme transport protein HutA: sequence of the gene, regulation of expression, and homology to the family of TonB-dependent proteins. J Bacteriol. 1994 Jun;176(11):3269–3277. doi: 10.1128/jb.176.11.3269-3277.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Henderson D. P., Payne S. M. Cloning and characterization of the Vibrio cholerae genes encoding the utilization of iron from haemin and haemoglobin. Mol Microbiol. 1993 Feb;7(3):461–469. doi: 10.1111/j.1365-2958.1993.tb01137.x. [DOI] [PubMed] [Google Scholar]
  27. Henderson D. P., Payne S. M. Vibrio cholerae iron transport systems: roles of heme and siderophore iron transport in virulence and identification of a gene associated with multiple iron transport systems. Infect Immun. 1994 Nov;62(11):5120–5125. doi: 10.1128/iai.62.11.5120-5125.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Hennecke H. Regulation of bacterial gene expression by metal-protein complexes. Mol Microbiol. 1990 Oct;4(10):1621–1628. doi: 10.1111/j.1365-2958.1990.tb00538.x. [DOI] [PubMed] [Google Scholar]
  29. Jarosik G. P., Sanders J. D., Cope L. D., Muller-Eberhard U., Hansen E. J. A functional tonB gene is required for both utilization of heme and virulence expression by Haemophilus influenzae type b. Infect Immun. 1994 Jun;62(6):2470–2477. doi: 10.1128/iai.62.6.2470-2477.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Karkhoff-Schweizer R. R., Schryvers A. B., Schweizer H. P. Cloning and sequence analysis of the fur gene encoding an iron-regulatory protein of Neisseria meningitidis. Gene. 1994 Apr 8;141(1):139–140. doi: 10.1016/0378-1119(94)90143-0. [DOI] [PubMed] [Google Scholar]
  31. Lee B. C., Hill P. Identification of an outer-membrane haemoglobin-binding protein in Neisseria meningitidis. J Gen Microbiol. 1992 Dec;138(12):2647–2656. doi: 10.1099/00221287-138-12-2647. [DOI] [PubMed] [Google Scholar]
  32. Lee B. C. Isolation and characterization of the haemin-binding proteins from Neisseria meningitidis. Microbiology. 1994 Jun;140(Pt 6):1473–1480. doi: 10.1099/00221287-140-6-1473. [DOI] [PubMed] [Google Scholar]
  33. Lee B. C. Isolation of an outer membrane hemin-binding protein of Haemophilus influenzae type b. Infect Immun. 1992 Mar;60(3):810–816. doi: 10.1128/iai.60.3.810-816.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Lee B. C. Isolation of haemin-binding proteins of Neisseria gonorrhoeae. J Med Microbiol. 1992 Feb;36(2):121–127. doi: 10.1099/00222615-36-2-121. [DOI] [PubMed] [Google Scholar]
  35. Lee B. C., Schryvers A. B. Specificity of the lactoferrin and transferrin receptors in Neisseria gonorrhoeae. Mol Microbiol. 1988 Nov;2(6):827–829. doi: 10.1111/j.1365-2958.1988.tb00095.x. [DOI] [PubMed] [Google Scholar]
  36. Lewis L. A., Dyer D. W. Identification of an iron-regulated outer membrane protein of Neisseria meningitidis involved in the utilization of hemoglobin complexed to haptoglobin. J Bacteriol. 1995 Mar;177(5):1299–1306. doi: 10.1128/jb.177.5.1299-1306.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Litwin C. M., Calderwood S. B. Role of iron in regulation of virulence genes. Clin Microbiol Rev. 1993 Apr;6(2):137–149. doi: 10.1128/cmr.6.2.137. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Létoffé S., Ghigo J. M., Wandersman C. Iron acquisition from heme and hemoglobin by a Serratia marcescens extracellular protein. Proc Natl Acad Sci U S A. 1994 Oct 11;91(21):9876–9880. doi: 10.1073/pnas.91.21.9876. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. McKenna W. R., Mickelsen P. A., Sparling P. F., Dyer D. W. Iron uptake from lactoferrin and transferrin by Neisseria gonorrhoeae. Infect Immun. 1988 Apr;56(4):785–791. doi: 10.1128/iai.56.4.785-791.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Mickelsen P. A., Blackman E., Sparling P. F. Ability of Neisseria gonorrhoeae, Neisseria meningitidis, and commensal Neisseria species to obtain iron from lactoferrin. Infect Immun. 1982 Mar;35(3):915–920. doi: 10.1128/iai.35.3.915-920.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Mickelsen P. A., Sparling P. F. Ability of Neisseria gonorrhoeae, Neisseria meningitidis, and commensal Neisseria species to obtain iron from transferrin and iron compounds. Infect Immun. 1981 Aug;33(2):555–564. doi: 10.1128/iai.33.2.555-564.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Mietzner T. A., Bolan G., Schoolnik G. K., Morse S. A. Purification and characterization of the major iron-regulated protein expressed by pathogenic Neisseriae. J Exp Med. 1987 Apr 1;165(4):1041–1057. doi: 10.1084/jem.165.4.1041. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Morse S. A., Chen C. Y., LeFaou A., Mietzner T. A. A potential role for the major iron-regulated protein expressed by pathogenic Neisseria species. Rev Infect Dis. 1988 Jul-Aug;10 (Suppl 2):S306–S310. doi: 10.1093/cid/10.supplement_2.s306. [DOI] [PubMed] [Google Scholar]
  44. Perry R. D. Acquisition and storage of inorganic iron and hemin by the yersiniae. Trends Microbiol. 1993 Jul;1(4):142–147. doi: 10.1016/0966-842x(93)90129-f. [DOI] [PubMed] [Google Scholar]
  45. Pettersson A., van der Ley P., Poolman J. T., Tommassen J. Molecular characterization of the 98-kilodalton iron-regulated outer membrane protein of Neisseria meningitidis. Infect Immun. 1993 Nov;61(11):4724–4733. doi: 10.1128/iai.61.11.4724-4733.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Postle K. TonB and the gram-negative dilemma. Mol Microbiol. 1990 Dec;4(12):2019–2025. doi: 10.1111/j.1365-2958.1990.tb00561.x. [DOI] [PubMed] [Google Scholar]
  47. Simonson C., Trivett T., DeVoe I. W. Energy-independent uptake of iron from citrate by isolated outer membranes of Neisseria meningitidis. Infect Immun. 1981 Feb;31(2):547–553. doi: 10.1128/iai.31.2.547-553.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Stojiljkovic I., Hantke K. Hemin uptake system of Yersinia enterocolitica: similarities with other TonB-dependent systems in gram-negative bacteria. EMBO J. 1992 Dec;11(12):4359–4367. doi: 10.1002/j.1460-2075.1992.tb05535.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Stojiljkovic I., Hantke K. Transport of haemin across the cytoplasmic membrane through a haemin-specific periplasmic binding-protein-dependent transport system in Yersinia enterocolitica. Mol Microbiol. 1994 Aug;13(4):719–732. doi: 10.1111/j.1365-2958.1994.tb00465.x. [DOI] [PubMed] [Google Scholar]
  50. Stojiljkovic I., Hwa V., de Saint Martin L., O'Gaora P., Nassif X., Heffron F., So M. The Neisseria meningitidis haemoglobin receptor: its role in iron utilization and virulence. Mol Microbiol. 1995 Feb;15(3):531–541. doi: 10.1111/j.1365-2958.1995.tb02266.x. [DOI] [PubMed] [Google Scholar]
  51. Thomas C. E., Sparling P. F. Identification and cloning of a fur homologue from Neisseria meningitidis. Mol Microbiol. 1994 Feb;11(4):725–737. doi: 10.1111/j.1365-2958.1994.tb00350.x. [DOI] [PubMed] [Google Scholar]
  52. Thompson S. A., Wang L. L., Sparling P. F. Cloning and nucleotide sequence of frpC, a second gene from Neisseria meningitidis encoding a protein similar to RTX cytotoxins. Mol Microbiol. 1993 Jul;9(1):85–96. doi: 10.1111/j.1365-2958.1993.tb01671.x. [DOI] [PubMed] [Google Scholar]
  53. Thompson S. A., Wang L. L., West A., Sparling P. F. Neisseria meningitidis produces iron-regulated proteins related to the RTX family of exoproteins. J Bacteriol. 1993 Feb;175(3):811–818. doi: 10.1128/jb.175.3.811-818.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Tsai J., Dyer D. W., Sparling P. F. Loss of transferrin receptor activity in Neisseria meningitidis correlates with inability to use transferrin as an iron source. Infect Immun. 1988 Dec;56(12):3132–3138. doi: 10.1128/iai.56.12.3132-3138.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Wooldridge K. G., Williams P. H. Iron uptake mechanisms of pathogenic bacteria. FEMS Microbiol Rev. 1993 Nov;12(4):325–348. doi: 10.1111/j.1574-6976.1993.tb00026.x. [DOI] [PubMed] [Google Scholar]
  56. van Putten J. P. Iron acquisition and the pathogenesis of meningococcal and gonococcal disease. Med Microbiol Immunol. 1990;179(6):289–295. doi: 10.1007/BF00189607. [DOI] [PubMed] [Google Scholar]

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