Abstract
Streptococcus pneumoniae is a causative agent of bacterial pneumonia, otitis media, meningitis, and bacteremia. It causes considerable morbidity and mortality throughout the world, especially among children, the elderly, and immunocompromised individuals. We have demonstrated previously that the growth of S. pneumoniae is limited under iron-depleted conditions and can be restored by the addition of either hemin or hemoglobin. In the present study, we showed that S. pneumoniae had the ability to bind hemin and that the level of hemin binding activity was not affected by supplementation of the growth medium with iron. Approximately 70 to 80% of the hemin binding activity was mediated by proteinase-resistant components, and the remainder was mediated by proteins. Hemin binding proteins were located in both soluble extract and envelope fractions of pneumococcal cells. By batch affinity chromatography, a major hemin binding polypeptide with an apparent molecular mass of 43 kDa was identified in the cell lysate of S. pneumoniae. Polyclonal antibodies against this polypeptide were raised. By immunoblot analysis, this hemin binding polypeptide was localized in the envelope and did not exhibit any variation in molecular weight among all serotypes tested. The subcellular distribution of hemin binding activity may have functional implications.
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- Adams T. J., Vartivarian S., Cowart R. E. Iron acquisition systems of Listeria monocytogenes. Infect Immun. 1990 Aug;58(8):2715–2718. doi: 10.1128/iai.58.8.2715-2718.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Adhikari P., Berish S. A., Nowalk A. J., Veraldi K. L., Morse S. A., Mietzner T. A. The fbpABC locus of Neisseria gonorrhoeae functions in the periplasm-to-cytosol transport of iron. J Bacteriol. 1996 Apr;178(7):2145–2149. doi: 10.1128/jb.178.7.2145-2149.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
- AlonsoDeVelasco E., Verheul A. F., Verhoef J., Snippe H. Streptococcus pneumoniae: virulence factors, pathogenesis, and vaccines. Microbiol Rev. 1995 Dec;59(4):591–603. doi: 10.1128/mr.59.4.591-603.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bagg A., Neilands J. B. Molecular mechanism of regulation of siderophore-mediated iron assimilation. Microbiol Rev. 1987 Dec;51(4):509–518. doi: 10.1128/mr.51.4.509-518.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Berry A. M., Yother J., Briles D. E., Hansman D., Paton J. C. Reduced virulence of a defined pneumolysin-negative mutant of Streptococcus pneumoniae. Infect Immun. 1989 Jul;57(7):2037–2042. doi: 10.1128/iai.57.7.2037-2042.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1016/0003-2697(76)90527-3. [DOI] [PubMed] [Google Scholar]
- Bramanti T. E., Holt S. C. Effect of porphyrins and host iron transport proteins on outer membrane protein expression in Porphyromonas (Bacteroides) gingivalis: identification of a novel 26 kDa hemin-repressible surface protein. Microb Pathog. 1992 Jul;13(1):61–73. doi: 10.1016/0882-4010(92)90032-j. [DOI] [PubMed] [Google Scholar]
- Carman R. J., Ramakrishnan M. D., Harper F. H. Hemin levels in culture medium of Porphyromonas (Bacteroides) gingivalis regulate both hemin binding and trypsinlike protease production. Infect Immun. 1990 Dec;58(12):4016–4019. doi: 10.1128/iai.58.12.4016-4019.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Crain M. J., Waltman W. D., 2nd, Turner J. S., Yother J., Talkington D. F., McDaniel L. S., Gray B. M., Briles D. E. Pneumococcal surface protein A (PspA) is serologically highly variable and is expressed by all clinically important capsular serotypes of Streptococcus pneumoniae. Infect Immun. 1990 Oct;58(10):3293–3299. doi: 10.1128/iai.58.10.3293-3299.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Crosa J. H. Genetics and molecular biology of siderophore-mediated iron transport in bacteria. Microbiol Rev. 1989 Dec;53(4):517–530. doi: 10.1128/mr.53.4.517-530.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cutler C. W., Eke P. I., Genco C. A., Van Dyke T. E., Arnold R. R. Hemin-induced modifications of the antigenicity and hemin-binding capacity of Porphyromonas gingivalis lipopolysaccharide. Infect Immun. 1996 Jun;64(6):2282–2287. doi: 10.1128/iai.64.6.2282-2287.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Daskaleros P. A., Payne S. M. Congo red binding phenotype is associated with hemin binding and increased infectivity of Shigella flexneri in the HeLa cell model. Infect Immun. 1987 Jun;55(6):1393–1398. doi: 10.1128/iai.55.6.1393-1398.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Desai P. J., Nzeribe R., Genco C. A. Binding and accumulation of hemin in Neisseria gonorrhoeae. Infect Immun. 1995 Dec;63(12):4634–4641. doi: 10.1128/iai.63.12.4634-4641.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Elkins C., Chen C. J., Thomas C. E. Characterization of the hgbA locus encoding a hemoglobin receptor from Haemophilus ducreyi. Infect Immun. 1995 Jun;63(6):2194–2200. doi: 10.1128/iai.63.6.2194-2200.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Evans S. L., Arceneaux J. E., Byers B. R., Martin M. E., Aranha H. Ferrous iron transport in Streptococcus mutans. J Bacteriol. 1986 Dec;168(3):1096–1099. doi: 10.1128/jb.168.3.1096-1099.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Genco C. A., Desai P. J. Iron acquisition in the pathogenic Neisseria. Trends Microbiol. 1996 May;4(5):179–184. doi: 10.1016/0966-842x(96)10029-9. [DOI] [PubMed] [Google Scholar]
- 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]
- 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]
- Kaca W., Roth R. I., Levin J. Hemoglobin, a newly recognized lipopolysaccharide (LPS)-binding protein that enhances LPS biological activity. J Biol Chem. 1994 Oct 7;269(40):25078–25084. [PubMed] [Google Scholar]
- Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
- 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]
- 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]
- 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]
- McDaniel L. S., Yother J., Vijayakumar M., McGarry L., Guild W. R., Briles D. E. Use of insertional inactivation to facilitate studies of biological properties of pneumococcal surface protein A (PspA). J Exp Med. 1987 Feb 1;165(2):381–394. doi: 10.1084/jem.165.2.381. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- 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]
- O'Connell W. A., Hickey E. K., Cianciotto N. P. A Legionella pneumophila gene that promotes hemin binding. Infect Immun. 1996 Mar;64(3):842–848. doi: 10.1128/iai.64.3.842-848.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Otto B. R., Sparrius M., Verweij-van Vught A. M., MacLaren D. M. Iron-regulated outer membrane protein of Bacteroides fragilis involved in heme uptake. Infect Immun. 1990 Dec;58(12):3954–3958. doi: 10.1128/iai.58.12.3954-3958.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Paton J. C., Andrew P. W., Boulnois G. J., Mitchell T. J. Molecular analysis of the pathogenicity of Streptococcus pneumoniae: the role of pneumococcal proteins. Annu Rev Microbiol. 1993;47:89–115. doi: 10.1146/annurev.mi.47.100193.000513. [DOI] [PubMed] [Google Scholar]
- Payne S. M. Iron acquisition in microbial pathogenesis. Trends Microbiol. 1993 May;1(2):66–69. doi: 10.1016/0966-842x(93)90036-q. [DOI] [PubMed] [Google Scholar]
- Payne S. M. Iron and virulence in the family Enterobacteriaceae. Crit Rev Microbiol. 1988;16(2):81–111. doi: 10.3109/10408418809104468. [DOI] [PubMed] [Google Scholar]
- 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]
- Perry R. D., Brubaker R. R. Accumulation of iron by yersiniae. J Bacteriol. 1979 Mar;137(3):1290–1298. doi: 10.1128/jb.137.3.1290-1298.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pidcock K. A., Wooten J. A., Daley B. A., Stull T. L. Iron acquisition by Haemophilus influenzae. Infect Immun. 1988 Apr;56(4):721–725. doi: 10.1128/iai.56.4.721-725.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Russell L. M., Cryz S. J., Jr, Holmes R. K. Genetic and biochemical evidence for a siderophore-dependent iron transport system in Corynebacterium diphtheriae. Infect Immun. 1984 Jul;45(1):143–149. doi: 10.1128/iai.45.1.143-149.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schneider R., Hantke K. Iron-hydroxamate uptake systems in Bacillus subtilis: identification of a lipoprotein as part of a binding protein-dependent transport system. Mol Microbiol. 1993 Apr;8(1):111–121. doi: 10.1111/j.1365-2958.1993.tb01208.x. [DOI] [PubMed] [Google Scholar]
- Schryvers A. B. Identification of the transferrin- and lactoferrin-binding proteins in Haemophilus influenzae. J Med Microbiol. 1989 Jun;29(2):121–130. doi: 10.1099/00222615-29-2-121. [DOI] [PubMed] [Google Scholar]
- Stoebner J. A., Payne S. M. Iron-regulated hemolysin production and utilization of heme and hemoglobin by Vibrio cholerae. Infect Immun. 1988 Nov;56(11):2891–2895. doi: 10.1128/iai.56.11.2891-2895.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Tai S. S., Lee C. J., Winter R. E. Hemin utilization is related to virulence of Streptococcus pneumoniae. Infect Immun. 1993 Dec;61(12):5401–5405. doi: 10.1128/iai.61.12.5401-5405.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Waltman W. D., McDaniel L. S., Gray B. M., Briles D. E. Variation in the molecular weight of PspA (pneumococcal surface protein A) among Streptococcus pneumoniae. Microb Pathog. 1990 Jan;8(1):61–69. doi: 10.1016/0882-4010(90)90008-e. [DOI] [PubMed] [Google Scholar]
- Worst D. J., Otto B. R., de Graaff J. Iron-repressible outer membrane proteins of Helicobacter pylori involved in heme uptake. Infect Immun. 1995 Oct;63(10):4161–4165. doi: 10.1128/iai.63.10.4161-4165.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Yother J., White J. M. Novel surface attachment mechanism of the Streptococcus pneumoniae protein PspA. J Bacteriol. 1994 May;176(10):2976–2985. doi: 10.1128/jb.176.10.2976-2985.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]