Home LiteratureArticle Details
PMID: 2187814 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

Calcium is required for binding of Escherichia coli hemolysin (HlyA) to erythrocyte membranes.

Infection and immunity ·Vol. 58 ·No. 6 ·1990-06-00 ·Pages 1951-8

Boehm DF, Welch RA, Snyder IS

Abstract

The calcium requirement for hemolytic activity of Escherichia coli hemolysin was investigated by using hemolytic assays and immunoblotting of toxin-treated erythrocytes. The hemolytic activity of cell culture supernatants obtained during growth of E. coli in Luria-Bertani (LB) broth or calcium-free LB broth was calcium dependent. The hemolytic activity of culture supernatants obtained during growth in LB broth supplemented with calcium was calcium independent. Osmotic protection experiments using Dextran 4 to prevent cell lysis indicated that calcium was required for the binding of hemolysin to erythrocytes at both 4 and 37 degrees C. The binding efficiency at 4 degrees C was 50% of that occurring at 37 degrees C. The calcium-dependent binding was confirmed by immunoblotting saline-washed, toxin-treated erythrocytes with a monoclonal antibody after sodium dodecyl sulfate-polyacrylamide gel electrophoresis separation of membrane proteins. Bound hemolysin increased the calcium permeability of the cell membranes as evidenced by calcium-induced membrane protein alterations. The alterations in membrane proteins did not directly cause lysis of the cells. The results were consistent with a mechanism of lysis involving the formation of cation-selective pores in the membranes of target cells.

MeSH Terms
Animals Bacterial Proteins/metabolism Calcium/metabolism Cell Membrane Permeability Erythrocyte Membrane/metabolism Escherichia coli/pathogenicity Escherichia coli Proteins Hemolysin Proteins Hemolysis Humans Immunoblotting Sheep
Chemicals
Bacterial Proteins Escherichia coli Proteins Hemolysin Proteins Hlya protein, E coli Calcium
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Boehm D F
Department of Microbiology and Immunology, West Virginia University Medical Center, Morgantown 26506.
Welch R A
Snyder I S
References (50)
50 references, click to expand
  1. Induction of histamine release from rat mast cells and human basophilic granulocytes by clinical Escherichia coli isolates and relation to hemolysin production and adhesin expression.
    J Clin Microbiol. 1988 Sep;26(9):1831-7 PMID: 2460497
  2. Alterations of amino acid repeats in the Escherichia coli hemolysin affect cytolytic activity and secretion.
    Proc Natl Acad Sci U S A. 1988 Jul;85(14):5269-73 PMID: 2839840
  3. Escherichia coli hemolysin may damage target cell membranes by generating transmembrane pores.
    Infect Immun. 1986 Apr;52(1):63-9 PMID: 3514465
  4. Cleavage of structural proteins during the assembly of the head of bacteriophage T4.
    Nature. 1970 Aug 15;227(5259):680-5 PMID: 5432063
  5. Membrane damage by pore-forming bacterial cytolysins.
    Microb Pathog. 1986 Feb;1(1):5-14 PMID: 2854594
  6. Synthesis and secretion of hemolysin by Escherichia coli.
    J Bacteriol. 1980 Oct;144(1):53-9 PMID: 6998968
  7. Electrophoretic analysis of the major polypeptides of the human erythrocyte membrane.
    Biochemistry. 1971 Jun 22;10(13):2606-17 PMID: 4326772
  8. Effect of Escherichia coli alpha-hemolysin on human peripheral leukocyte function in vitro.
    Infect Immun. 1982 Sep;37(3):966-74 PMID: 6752033
  9. Chemical and immunological analysis of the complex structure of Escherichia coli alpha-hemolysin.
    J Bacteriol. 1985 Dec;164(3):1071-80 PMID: 3905764
  10. Effect of Escherichia coli hemolysin on permeability of erythrocyte membranes to calcium.
    Toxicon. 1986;24(6):559-66 PMID: 3750344
  11. Mutations affecting activity and transport of haemolysin in Escherichia coli.
    Mol Gen Genet. 1987 Feb;206(2):238-45 PMID: 3295483
  12. The carboxy-terminal region of haemolysin 2001 is required for secretion of the toxin from Escherichia coli.
    Mol Gen Genet. 1986 Oct;205(1):127-33 PMID: 3025555
  13. In vitro cytotoxic effect of alpha-hemolytic and nonhemolytic Escherichia coli on human blood granulocytes and monocytes.
    Acta Pathol Microbiol Immunol Scand B. 1987 Aug;95(4):219-25 PMID: 3314335
  14. Escherichia coli hemolysin is released extracellularly without cleavage of a signal peptide.
    J Bacteriol. 1985 Jul;163(1):88-93 PMID: 3891742
  15. The preparation and chemical characteristics of hemoglobin-free ghosts of human erythrocytes.
    Arch Biochem Biophys. 1963 Jan;100:119-30 PMID: 14028302
  16. Association of hemolysin production, hemagglutination of human erythrocytes, and virulence for chicken embryos of extraintestinal Escherichia coli isolates.
    Infect Immun. 1978 Apr;20(1):50-4 PMID: 352937
  17. Cytotoxic activity of partially purified Escherichia coli alpha haemolysin.
    J Med Microbiol. 1982 Feb;15(1):11-21 PMID: 6815328
  18. Escherichia coli hemolysin permeabilizes small unilamellar vesicles loaded with calcein by a single-hit mechanism.
    FEBS Lett. 1988 May 9;232(1):217-20 PMID: 3284764
  19. Characterization of Escherichia coli hemolysins conferring quantitative differences in virulence.
    Infect Immun. 1984 Jan;43(1):156-60 PMID: 6317564
  20. Pore formation by the Escherichia coli hemolysin: evidence for an association-dissociation equilibrium of the pore-forming aggregates.
    Infect Immun. 1989 Mar;57(3):887-95 PMID: 2465272
  21. Regulation of haemolysin synthesis in E. coli determined by HLY genes of human origin.
    Mol Gen Genet. 1985;199(1):111-6 PMID: 3923298
  22. Properties of strains of Escherichia coli isolated from a variety of sources.
    J Med Microbiol. 1975 Feb;8(1):107-11 PMID: 1092859
  23. The kinetics of erythrocyte lysis by Escherichia coli haemolysin.
    J Med Microbiol. 1974 May;7(2):189-95 PMID: 4134504
  24. Formation of gamma-glutamyl-epsilon-lysine bridges between membrane proteins by a Ca2+-regulated enzyme in intact erythrocytes.
    J Supramol Struct. 1978;9(3):427-40 PMID: 34754
  25. Damage to cell membranes by pore-forming bacterial cytolysins.
    Prog Allergy. 1988;40:1-43 PMID: 2451254
  26. Some factors affecting production and assay of Escherichia coli haemolysins.
    J Gen Microbiol. 1969 Jan;55(1):139-43 PMID: 4886934
  27. Effects of intracellular Ca2+ and proteolytic digestion of the membrane skeleton on the mechanical properties of the red blood cell membrane.
    Biochim Biophys Acta. 1987 Nov 27;905(1):181-94 PMID: 2445380
  28. Release of a chimeric protein into the medium from Escherichia coli using the C-terminal secretion signal of haemolysin.
    EMBO J. 1987 Sep;6(9):2835-41 PMID: 3119330
  29. The haemolytic activity of Bact. coli.
    J Pathol Bacteriol. 1952 Oct;64(4):751-61 PMID: 13000589
  30. Limited proteolysis of the erythrocyte membrane skeleton by calcium-dependent proteinases.
    Biochim Biophys Acta. 1986 Jul 16;882(3):287-96 PMID: 3015225
  31. Secretion of haemolysin by Escherichia coli.
    Curr Top Microbiol Immunol. 1986;125:159-81 PMID: 3017638
  32. The toxic role of alpha-haemolysin in the pathogenesis of experimental Escherichia coli infection in mice.
    J Gen Microbiol. 1985 Feb;131(2):395-403 PMID: 3884734
  33. Membrane damage by hemolytic viruses, toxins, complement, and other cytotoxic agents. A common mechanism blocked by divalent cations.
    J Biol Chem. 1986 Jul 15;261(20):9300-8 PMID: 3013883
  34. Potent leukocidal action of Escherichia coli hemolysin mediated by permeabilization of target cell membranes.
    J Exp Med. 1989 Mar 1;169(3):737-54 PMID: 2538544
  35. Haemolysin contributes to virulence of extra-intestinal E. coli infections.
    Nature. 1981 Dec 17;294(5842):665-7 PMID: 7031483
  36. Calcium effects on human erythrocyte membrane proteins.
    Biochim Biophys Acta. 1977 Nov 15;471(1):162-8 PMID: 921972
  37. Properties of the Hemolytic Activities of Escherichia coli.
    Infect Immun. 1971 May;3(5):678-87 PMID: 16558036
  38. In vivo function of hemolysin in the nephropathogenicity of Escherichia coli.
    Infect Immun. 1983 Oct;42(1):245-9 PMID: 6352494
  39. Effect of Escherichia coli alpha-hemolysin on human peripheral leukocyte viability in vitro.
    Infect Immun. 1982 May;36(2):455-61 PMID: 7044971
  40. Calcium accumulation in human and sheep erythrocytes that is induced by Escherichia coli hemolysin.
    Toxicon. 1983;21(5):717-27 PMID: 6359585
  41. Transcriptional organization of the Escherichia coli hemolysin genes.
    J Bacteriol. 1988 Apr;170(4):1622-30 PMID: 2450867
  42. Quantitative study of the binding and hemolytic efficiency of Escherichia coli hemolysin.
    Infect Immun. 1989 Mar;57(3):983-8 PMID: 2465275
  43. Calcium-induced erythrocyte membrane changes. The role of adsorption of cytosol proteins and proteases.
    Biochim Biophys Acta. 1979 Feb 20;551(1):1-9 PMID: 427145
  44. Nucleotide sequence of an Escherichia coli chromosomal hemolysin.
    J Bacteriol. 1985 Jul;163(1):94-105 PMID: 3891743
  45. The repeat domain of Escherichia coli haemolysin (HlyA) is responsible for its Ca2+-dependent binding to erythrocytes.
    Mol Gen Genet. 1988 Nov;214(3):553-61 PMID: 3063951
  46. The haemolysins of Escherichia coli.
    J Pathol Bacteriol. 1963 Jan;85:197-211 PMID: 13989441
  47. Escherichia coli haemolysin forms voltage-dependent ion channels in lipid membranes.
    Biochim Biophys Acta. 1987 Nov 27;905(1):109-17 PMID: 2445378
  48. The role of alpha haemolysin in the virulence of Escherichia coli for mice.
    J Med Microbiol. 1982 Feb;15(1):23-30 PMID: 6754940
  49. Mechanism of Escherichia coli alpha-hemolysin-induced injury to isolated renal tubular cells.
    Am J Pathol. 1987 Feb;126(2):350-7 PMID: 3030115
  50. Improved staining of proteins in polyacrylamide gels including isoelectric focusing gels with clear background at nanogram sensitivity using Coomassie Brilliant Blue G-250 and R-250.
    Electrophoresis. 1988 Jun;9(6):255-62 PMID: 2466658
Article Info
Journal
Infection and immunity
Abbr.
Infect Immun
ISSN
0019-9567
DOI
10.1128/iai.58.6.1951-1958.1990
Published
1990-06-00
Pages
1951-8
Language
English
Region
United States
NLM ID
0246127
PMCID
PMC258749
Subset
IM
Grants
NIDDK NIH HHS · R01 DK063250 · United States
NIAID NIH HHS · AI-20323 · United States
External Links
PubMed source DOI: 10.1128/iai.58.6.1951-1958.1990 PMC Full Text View journal details
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: product@genelibs.com

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2