FREE SHIPPING over $40
OVER 9000 FIVE-STAR REVIEWS
5% OFF ALL SUBSCRIPTIONS
100% HAPPINESS GUARANTEE
Analyze Diet
0
Home / Equine Research Bank / Infect Immun / Cloning, expression, sequence analysis, and characterization...
Infection and immunity1999; 67(12); 6478-6486; doi: 10.1128/IAI.67.12.6478-6486.1999

Cloning, expression, sequence analysis, and characterization of streptokinases secreted by porcine and equine isolates of Streptococcus equisimilis.

Abstract: Streptokinases secreted by nonhuman isolates of group C streptococci (Streptococcus equi, S. equisimilis, and S. zooepidemicus) have been shown to bind to different mammalian plasminogens but exhibit preferential plasminogen activity. The streptokinase genes from S. equisimilis strains which activated either equine or porcine plasminogen were cloned, sequenced, and expressed in Escherichia coli. The streptokinase secreted by the equine isolate had little similarity to any known streptokinases secreted by either human or porcine isolates. The streptokinase secreted by the porcine isolate had limited structural and functional similarities to streptokinases secreted by human isolates. Plasminogen activation studies with immobilized (His)(6)-tagged recombinant streptokinases indicated that these recombinant streptokinases interacted with plasminogen in a manner similar to that observed when streptokinase and plasminogen interact in the fluid phase. Analysis of the cleavage products of the streptokinase-plasminogen interaction indicated that human, equine, and porcine plasminogens were all cleaved at the same highly conserved site. The site at which streptokinase was cleaved to form altered streptokinase (Sk*) was also determined. This study confirmed not only the presence of streptokinases in nonhuman S. equisimilis isolates but also that these proteins belong to a family of plasminogen activators more diverse than previously thought.
Get Full Text
Publication Date: 1999-11-24 PubMed ID: 10569766PubMed Central: PMC97058DOI: 10.1128/IAI.67.12.6478-6486.1999Google Scholar: Lookup
The Equine Research Bank provides access to a large database of publicly available scientific literature. Inclusion in the Research Bank does not imply endorsement of study methods or findings by Mad Barn.
LinkedInCopy
  • Journal Article
  • Research Support
  • Non-U.S. Gov't
  • Research Support
  • U.S. Gov't
  • P.H.S.

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This research involves cloning, sequencing, and studying the similarity and differences of streptokinases, specifically those produced by porcine and equine isolates of Streptococcus equisimilis. The findings suggest that these streptokinases cause plasminogen activity and resemble each other in some ways, but they also show some significant differences from known streptokinases.

Streptokinases and Plasminogen Activation

  • The study mentions streptokinases, enzymes produced by various types of streptococci bacteria, including group C species like S. equi, S. equisimilis, and S. zooepidemicus.
  • These streptokinases have been found to bind to different mammals’ plasminogens and demonstrate preferential plasminogen activity, implying they foster the conversion of plasminogen into plasmin, a crucial process in the breakdown of blood clots.

Cloning and Sequencing of Streptokinase Genes

  • The research involved cloning and sequencing streptokinase genes from S. equisimilis strains known to activate either horse or pig plasminogen.
  • The resulting sequenced genes were then expressed in Escherichia coli, a common bacteria often used in genetic research due to its rapid growth and well-understood genetics.

Comparative Analysis of Streptokinases

  • The streptokinase secreted by equine isolate showed minimal similarity to known streptokinases from human and pig isolates. The discrepancy might imply distinct evolutionary paths or functional mechanisms among these enzymes.
  • In contrast, the streptokinase produced by the pig isolate had some structural and functional resemblance to known human streptokinases, although these similarities were limited.

Plasminogen Interaction with Recombinant Streptokinases

  • Experiments with immobilized (His)(6)-tagged recombinant streptokinases pointed out that these interacted with plasminogen similarly to how streptokinase and plasminogen interact in a fluid phase.
  • The research found that all three types of plasminogen (human, horse, and pig) were cleaved at the same highly conserved position from their interaction with the streptokinase.

Broader Implications

  • This study confirms the presence of streptokinases in nonhuman S. equisimilis isolates and reveals that these proteins belong to a group of plasminogen activators more diverse than previously thought.
  • This discovery has significant implications for understanding the physiological roles of streptokinases, their potential therapeutic uses, and the broader mechanisms of plasminogen activation and fibrinolysis in different species.

Cite This Article

APA
Caballero AR, Lottenberg R, Johnston KH. (1999). Cloning, expression, sequence analysis, and characterization of streptokinases secreted by porcine and equine isolates of Streptococcus equisimilis. Infect Immun, 67(12), 6478-6486. https://doi.org/10.1128/IAI.67.12.6478-6486.1999

Publication

Infection and immunity
ISSN: 0019-9567
NlmUniqueID: 0246127
Country: United States
Language: English
Volume: 67
Issue: 12
Pages: 6478-6486

Researcher Affiliations

Caballero, A R
  • Department of Microbiology, Immunology and Parasitology, Louisiana State University Medical Center, New Orleans, Louisiana 70112, USA.
Lottenberg, R
    Johnston, K H

      MeSH Terms

      • Amino Acid Sequence
      • Animals
      • Base Sequence
      • Cloning, Molecular
      • Horse Diseases / microbiology
      • Horses
      • Humans
      • Molecular Sequence Data
      • Plasminogen / isolation & purification
      • Plasminogen / metabolism
      • Plasminogen Activators / metabolism
      • Polymerase Chain Reaction
      • Sequence Analysis, DNA
      • Streptococcal Infections / microbiology
      • Streptococcal Infections / veterinary
      • Streptococcus / enzymology
      • Streptococcus / genetics
      • Streptokinase / chemistry
      • Streptokinase / genetics
      • Streptokinase / metabolism
      • Swine
      • Swine Diseases / microbiology

      Grant Funding

      • R01DK45014 / NIDDK NIH HHS

      References

      This article includes 39 references
      1. Arditi M, Shulman ST, Davis AT, Yogev R. Group C beta-hemolytic streptococcal infections in children: nine pediatric cases and review.. Rev Infect Dis 1989 Jan-Feb;11(1):34-45.
        pubmed: 2644689doi: 10.1093/clinids/11.1.34google scholar: lookup
      2. Ausubel FM, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K. Current protocols in molecular biology. New York, N.Y: Wiley Interscience; 1993.
      3. Boyle MD, Lottenberg R. Plasminogen activation by invasive human pathogens.. Thromb Haemost 1997 Jan;77(1):1-10.
        pubmed: 9031440
      4. Brockway WJ, Castellino FJ. A characterization of native streptokinase and altered streptokinase isolated from a human plasminogen activator complex.. Biochemistry 1974 May 7;13(10):2063-70.
        pubmed: 4857058doi: 10.1021/bi00707a010google scholar: lookup
      5. Deutsch DG, Mertz ET. Plasminogen: purification from human plasma by affinity chromatography.. Science 1970 Dec 4;170(3962):1095-6.
        pubmed: 5475635doi: 10.1126/science.170.3962.1095google scholar: lookup
      6. Diebel RH, Steeley WH. Streptococcacea. Bergey's manual of determinative bacteriology Baltimore, Md: Williams and Wilkins; 1974. pp. 490–517.
      7. Dunn SD. Effects of the modification of transfer buffer composition and the renaturation of proteins in gels on the recognition of proteins on Western blots by monoclonal antibodies.. Anal Biochem 1986 Aug 15;157(1):144-53.
        pubmed: 3532863doi: 10.1016/0003-2697(86)90207-1google scholar: lookup
      8. Forsgren M, Råden B, Israelsson M, Larsson K, Hedén LO. Molecular cloning and characterization of a full-length cDNA clone for human plasminogen.. FEBS Lett 1987 Mar 23;213(2):254-60.
        pubmed: 3030813doi: 10.1016/0014-5793(87)81501-6google scholar: lookup
      9. Holm SE, Bergholm AM, Johnston KH. A streptococcal plasminogen activator in the focus of infection and in the kidneys during the initial phase of experimental streptococcal glomerulonephritis.. APMIS 1988 Dec;96(12):1097-108.
        pubmed: 3063303doi: 10.1111/j.1699-0463.1988.tb00987.xgoogle scholar: lookup
      10. Huang TT, Malke H, Ferretti JJ. The streptokinase gene of group A streptococci: cloning, expression in Escherichia coli, and sequence analysis.. Mol Microbiol 1989 Feb;3(2):197-205.
        pubmed: 2668686doi: 10.1111/j.1365-2958.1989.tb01808.xgoogle scholar: lookup
      11. Johnsen LB, Poulsen K, Kilian M, Petersen TE. Purification and cloning of a streptokinase from Streptococcus uberis.. Infect Immun 1999 Mar;67(3):1072-8.
        pmc: PMC96431pubmed: 10024545doi: 10.1128/iai.67.3.1072-1078.1999google scholar: lookup
      12. Johnston KH. Solid and fluid phase assays for bacterial plasminogen activators. J Microbiol Methods 1993;18:267–274.
      13. Johnston KH, Chaiban JE, Wheeler RC. Analysis of the variable domain of the streptokinase gene from streptococci associated with post-streptococcal glomerulonephritis. Zentbl Bakteriol Suppl 1992;22:339–342.
      14. Johnston KH, Zabriskie JB. Purification and partial characterization of the nephritis strain-associated protein from Streptococcus pyogenes, group A.. J Exp Med 1986 Mar 1;163(3):697-712.
        pmc: PMC2188047pubmed: 3512759doi: 10.1084/jem.163.3.697google scholar: lookup
      15. Kohn J, Wilchek M. A new approach (cyano-transfer) for cyanogen bromide activation of Sepharose at neutral pH, which yields activated resins, free of interfering nitrogen derivatives.. Biochem Biophys Res Commun 1982 Aug;107(3):878-84.
        pubmed: 7138526doi: 10.1016/0006-291x(82)90604-0google scholar: lookup
      16. Kulisek ES, Holm SE, Johnston KH. A chromogenic assay for the detection of plasmin generated by plasminogen activator immobilized on nitrocellulose using a para-nitroanilide synthetic peptide substrate.. Anal Biochem 1989 Feb 15;177(1):78-84.
        pubmed: 2525884doi: 10.1016/0003-2697(89)90017-1google scholar: lookup
      17. Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4.. Nature 1970 Aug 15;227(5259):680-5.
        pubmed: 5432063doi: 10.1038/227680a0google scholar: lookup
      18. Leigh JA. Purification of a plasminogen activator from Streptococcus uberis.. FEMS Microbiol Lett 1994 May 1;118(1-2):153-8.
        pubmed: 8013872doi: 10.1111/j.1574-6968.1994.tb06818.xgoogle scholar: lookup
      19. Leigh JA, Hodgkinson SM, Lincoln RA. The interaction of Streptococcus dysgalactiae with plasmin and plasminogen.. Vet Microbiol 1998 Mar 15;61(1-2):121-35.
        pubmed: 9646471doi: 10.1016/s0378-1135(98)00179-5google scholar: lookup
      20. Lizano S, Johnston KH. Streptokinase-mediated plasminogen activation using a recombinant dual fusion protein construct. A novel approach to study bacterial-host protein interactions. J Microbiol Methods 1995;23:261–280.
      21. Lottenberg R, Broder CC, Boyle MD, Kain SJ, Schroeder BL, Curtiss R 3rd. Cloning, sequence analysis, and expression in Escherichia coli of a streptococcal plasmin receptor.. J Bacteriol 1992 Aug;174(16):5204-10.
        pmc: PMC206353pubmed: 1322883doi: 10.1128/jb.174.16.5204-5210.1992google scholar: lookup
      22. Malke H. Polymorphism of the streptokinase gene: implications for the pathogenesis of post-streptococcal glomerulonephritis.. Zentralbl Bakteriol 1993 Apr;278(2-3):246-57.
        pubmed: 8347929doi: 10.1016/s0934-8840(11)80842-xgoogle scholar: lookup
      23. Malke H, Ferretti JJ. Streptokinase: cloning, expression, and excretion by Escherichia coli.. Proc Natl Acad Sci U S A 1984 Jun;81(11):3557-61.
        pmc: PMC345548pubmed: 6374659doi: 10.1073/pnas.81.11.3557google scholar: lookup
      24. Malke H, Roe B, Ferretti JJ. Nucleotide sequence of the streptokinase gene from Streptococcus equisimilis H46A.. Gene 1985;34(2-3):357-62.
        pubmed: 2989113doi: 10.1016/0378-1119(85)90145-3google scholar: lookup
      25. Marti T, Schaller J, Rickli EE. Determination of the complete amino-acid sequence of porcine miniplasminogen.. Eur J Biochem 1985 Jun 3;149(2):279-85.
        pubmed: 3846533doi: 10.1111/j.1432-1033.1985.tb08923.xgoogle scholar: lookup
      26. McCoy HE, Broder CC, Lottenberg R. Streptokinases produced by pathogenic group C streptococci demonstrate species-specific plasminogen activation.. J Infect Dis 1991 Sep;164(3):515-21.
        pubmed: 1869838doi: 10.1093/infdis/164.3.515google scholar: lookup
      27. Monsen JJ, Holm SE, Burman LG. A general method for cell lysis and preparation of DNA from streptococci. FEMS Microbiol Lett 1983;16:19–24.
      28. Myers EW, Miller W. Optimal alignments in linear space.. Comput Appl Biosci 1988 Mar;4(1):11-7.
        pubmed: 3382986doi: 10.1093/bioinformatics/4.1.11google scholar: lookup
      29. Needleman SB, Wunsch CD. A general method applicable to the search for similarities in the amino acid sequence of two proteins.. J Mol Biol 1970 Mar;48(3):443-53.
        pubmed: 5420325doi: 10.1016/0022-2836(70)90057-4google scholar: lookup
      30. Nordstrand A, Norgren M, Ferretti JJ, Holm SE. Streptokinase as a mediator of acute post-streptococcal glomerulonephritis in an experimental mouse model.. Infect Immun 1998 Jan;66(1):315-21.
        pmc: PMC107892pubmed: 9423873doi: 10.1128/iai.66.1.315-321.1998google scholar: lookup
      31. Nowicki ST, Minning-Wenz D, Johnston KH, Lottenberg R. Characterization of a novel streptokinase produced by Streptococcus equisimilis of non-human origin.. Thromb Haemost 1994 Oct;72(4):595-603.
        pubmed: 7878639
      32. Ohkuni H, Todome Y, Suzuki H, Mizuse M, Kotani N, Horiuchi K, Shikama N, Tsugita A, Johnston KH. Immunochemical studies and complete amino acid sequence of the streptokinase from Streptococcus pyogenes (group A) M type 12 strain A374.. Infect Immun 1992 Jan;60(1):278-83.
        pmc: PMC257533pubmed: 1370275doi: 10.1128/iai.60.1.278-283.1992google scholar: lookup
      33. Reddy KN. Streptokinase--biochemistry and clinical application.. Enzyme 1988;40(2-3):79-89.
        pubmed: 3049067doi: 10.1159/000469149google scholar: lookup
      34. Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors.. Proc Natl Acad Sci U S A 1977 Dec;74(12):5463-7.
        pmc: PMC431765pubmed: 271968doi: 10.1073/pnas.74.12.5463google scholar: lookup
      35. Schaller J, Straub C, Kämpfer U, Rickli EE. Complete amino acid sequence of equine miniplasminogen.. Protein Seq Data Anal 1991 Aug;4(2):69-74.
        pubmed: 1946332
      36. Smith PK, Krohn RI, Hermanson GT, Mallia AK, Gartner FH, Provenzano MD, Fujimoto EK, Goeke NM, Olson BJ, Klenk DC. Measurement of protein using bicinchoninic acid.. Anal Biochem 1985 Oct;150(1):76-85.
        pubmed: 3843705doi: 10.1016/0003-2697(85)90442-7google scholar: lookup
      37. Walter F, Siegel M, Malke H. Nucleotide sequence of the streptokinase gene from a Streptococcus pyogenes type 1 strain.. Nucleic Acids Res 1989 Feb 11;17(3):1261.
        pmc: PMC331761pubmed: 2646590doi: 10.1093/nar/17.3.1261google scholar: lookup
      38. Walter F, Siegel M, Malke H. Nucleotide sequence of the streptokinase gene from a group-G Streptococcus.. Nucleic Acids Res 1989 Feb 11;17(3):1262.
        pmc: PMC331762pubmed: 2922269doi: 10.1093/nar/17.3.1262google scholar: lookup
      39. Wang X, Lin X, Loy JA, Tang J, Zhang XC. Crystal structure of the catalytic domain of human plasmin complexed with streptokinase.. Science 1998 Sep 11;281(5383):1662-5.
        pubmed: 9733510doi: 10.1126/science.281.5383.1662google scholar: lookup

      Citations

      This article has been cited 9 times.
      1. Oh SI, Kim JW, Jung JY, Chae M, Lee YR, Kim JH, So B, Kim HY. Pathologic and molecular characterization of Streptococcus dysgalactiae subsp. equisimilis infection in neonatal piglets.. J Vet Sci 2018 Mar 31;19(2):313-317.
        doi: 10.4142/jvs.2018.19.2.313pubmed: 29284213google scholar: lookup
      2. Pinho MD, Erol E, Ribeiro-Gonçalves B, Mendes CI, Carriço JA, Matos SC, Preziuso S, Luebke-Becker A, Wieler LH, Melo-Cristino J, Ramirez M. Beta-hemolytic Streptococcus dysgalactiae strains isolated from horses are a genetically distinct population within the Streptococcus dysgalactiae taxon.. Sci Rep 2016 Aug 17;6:31736.
        doi: 10.1038/srep31736pubmed: 27530432google scholar: lookup
      3. Ciszewski M, Zegarski K, Szewczyk EM. Streptococcus dysgalactiae subsp. equisimilis Isolated From Infections in Dogs and Humans: Are Current Subspecies Identification Criteria accurate?. Curr Microbiol 2016 Nov;73(5):684-688.
        doi: 10.1007/s00284-016-1113-xpubmed: 27502064google scholar: lookup
      4. Kasuya K, Yoshida E, Harada R, Hasegawa M, Osaka H, Kato M, Shibahara T. Systemic Streptococcus dysgalactiae subspecies equisimilis infection in a Yorkshire pig with severe disseminated suppurative meningoencephalomyelitis.. J Vet Med Sci 2014 May;76(5):715-8.
        doi: 10.1292/jvms.13-0526pubmed: 24419876google scholar: lookup
      5. Egan SA, Ward PN, Watson M, Field TR, Leigh JA. Vru (S뀔4) controls expression of proven and putative virulence determinants and alters the ability of Streptococcus uberis to cause disease in dairy cattle.. Microbiology (Reading) 2012 Jun;158(Pt 6):1581-1592.
        doi: 10.1099/mic.0.055863-0pubmed: 22383474google scholar: lookup
      6. Jensen A, Kilian M. Delineation of Streptococcus dysgalactiae, its subspecies, and its clinical and phylogenetic relationship to Streptococcus pyogenes.. J Clin Microbiol 2012 Jan;50(1):113-26.
        doi: 10.1128/JCM.05900-11pubmed: 22075580google scholar: lookup
      7. Preziuso S, Laus F, Tejeda AR, Valente C, Cuteri V. Detection of Streptococcus dysgalactiae subsp. equisimilis in equine nasopharyngeal swabs by PCR.. J Vet Sci 2010 Mar;11(1):67-72.
        doi: 10.4142/jvs.2010.11.1.67pubmed: 20195067google scholar: lookup
      8. Ward PN, Field TR, Rapier CD, Leigh JA. The activation of bovine plasminogen by PauA is not required for virulence of Streptococcus uberis.. Infect Immun 2003 Dec;71(12):7193-6.
        doi: 10.1128/IAI.71.12.7193-7196.2003pubmed: 14638815google scholar: lookup
      9. Ward PN, Leigh JA. Characterization of PauB, a novel broad-spectrum plasminogen activator from Streptococcus uberis.. J Bacteriol 2002 Jan;184(1):119-25.
        doi: 10.1128/JB.184.1.119-125.2002pubmed: 11741851google scholar: lookup
      Subscribe to our newsletter
      Join 99,357 horse owners like you who receive our email updates on horse health and nutrition.