Skip to main content
NCBI home page
Biochemical Journal logoLink to Biochemical Journal
. 1990 Oct 1;271(1):87–91. doi: 10.1042/bj2710087

Retrograde axonal transport of an exogenous enzyme covalently linked to B-IIb fragment of tetanus toxin.

P Beaude 1, A Delacour 1, B Bizzini 1, D Domuado 1, M H Remy 1
PMCID: PMC1149516  PMID: 1699518

Abstract

Attempt to replace enzymes in a number of fatal lysosomal storage disease involving the central nervous system have as yet been unsuccessful owing to the impermeability of the blood/brain barrier to macromolecules. In order to treat storage disease due to enzyme deficiencies, we investigated the feasibility of transporting an enzyme into the central nervous system without crossing the blood/brain barrier. Using the B-IIb fragment of tetanus toxin (because it is involved in recognition by the nerve-cell endings), retrograde axonal transport toward the spinal cord and trans-synaptic movement, and glucose oxidase as a marker, we demonstrated that a non-toxic enzyme-vector conjugate was taken up by axon terminals. After injection into the gastrocnemius muscle, the B-IIb-glucose oxidase conjugate was detected, both histologically and electrochemically, distally to a ligature on the sciatic nerve. Thus the B-IIb fragment could serve as a vector for glucose oxidase transport into the central nervous system. It was also verified that the transported enzyme retained its activity. Transport of this 150 kDa molecule by fragment B-IIb of tetanus toxin suggests that other enzymes of a lesser molecular mass may also be transported.

Full text

PDF
87

Images in this article

89Fig. 2.
on p.89
89Fig. 3.
on p.89

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aubrée-Lecat A., Hervagault C., Delacour A., Beaude P., Bourdillon C., Remy M. H. Direct electrochemical determination of glucose oxidase in biological samples. Anal Biochem. 1989 May 1;178(2):427–430. doi: 10.1016/0003-2697(89)90665-9. [DOI] [PubMed] [Google Scholar]
  2. Bizzini B., Grob P., Akert K. Papain-derived fragment IIc of tetanus toxin: its binding to isolated synaptic membranes and retrograde axonal transport. Brain Res. 1981 Apr 6;210(1-2):291–299. doi: 10.1016/0006-8993(81)90902-1. [DOI] [PubMed] [Google Scholar]
  3. Bizzini B., Grob P., Glicksman M. A., Akert K. Use of the B-IIb tetanus toxin derived fragment as a specific neuropharmacological transport agent. Brain Res. 1980 Jul 7;193(1):221–227. doi: 10.1016/0006-8993(80)90959-2. [DOI] [PubMed] [Google Scholar]
  4. Bizzini B., Stoeckel K., Schwab M. An antigenic polypeptide fragment isolated from tetanus toxin: chemical characterization, binding to gangliosides and retrograde axonal transport in various neuron systems. J Neurochem. 1977 Mar;28(3):529–542. doi: 10.1111/j.1471-4159.1977.tb10423.x. [DOI] [PubMed] [Google Scholar]
  5. Büttner-Ennever J. A., Grob P., Akert K., Bizzini B. Transsynaptic retrograde labeling in the oculomotor system of the monkey with [125I]tetanus toxin BIIb fragment. Neurosci Lett. 1981 Nov 4;26(3):233–238. doi: 10.1016/0304-3940(81)90138-5. [DOI] [PubMed] [Google Scholar]
  6. Carlsson J., Drevin H., Axén R. Protein thiolation and reversible protein-protein conjugation. N-Succinimidyl 3-(2-pyridyldithio)propionate, a new heterobifunctional reagent. Biochem J. 1978 Sep 1;173(3):723–737. doi: 10.1042/bj1730723. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Demignot S., Domurado D. Effect of prosthetic sugar groups on the pharmacokinetics of glucose-oxidase. Drug Des Deliv. 1987 May;1(4):333–348. [PubMed] [Google Scholar]
  8. Dimpfel W., Huang R. T., Habermann E. Gangliosides in nervous tissue cultures and binding of 125I-labelled tetanus toxin, a neuronal marker. J Neurochem. 1977 Aug;29(2):329–334. doi: 10.1111/j.1471-4159.1977.tb09626.x. [DOI] [PubMed] [Google Scholar]
  9. Dumas M., Schwab M. E., Baumann R., Thoenen H. Retrograde transport of tetanus toxin through a chain of two neurons. Brain Res. 1979 Apr 13;165(2):354–357. doi: 10.1016/0006-8993(79)90569-9. [DOI] [PubMed] [Google Scholar]
  10. Dumas M., Schwab M. E., Thoenen H. Retrograde axonal transport of specific macromolecules as a tool for characterizing nerve terminal membranes. J Neurobiol. 1979 Mar;10(2):179–197. doi: 10.1002/neu.480100207. [DOI] [PubMed] [Google Scholar]
  11. Evinger C., Erichsen J. T. Transsynaptic retrograde transport of fragment C of tetanus toxin demonstrated by immunohistochemical localization. Brain Res. 1986 Aug 20;380(2):383–388. doi: 10.1016/0006-8993(86)90241-6. [DOI] [PubMed] [Google Scholar]
  12. Fairweather N. F., Lyness V. A., Pickard D. J., Allen G., Thomson R. O. Cloning, nucleotide sequencing, and expression of tetanus toxin fragment C in Escherichia coli. J Bacteriol. 1986 Jan;165(1):21–27. doi: 10.1128/jb.165.1.21-27.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Fedinec A. A., Toth P., Bizzini B. Relation of spastic and flaccid paralysis to retrograde transport of 125I-tetanus toxin and its 125I-Ibc fragment. Modulating effect of F (ab) antibodies directed to specific areas on the toxin molecule. Boll Ist Sieroter Milan. 1985;64(1):35–41. [PubMed] [Google Scholar]
  14. Fillenz M., Gagnon C., Stoeckel K., Thoenen H. Selective uptake and retrograde axonal transport of dopamine-beta-hydroxylase antibodies in peripheral adrenergic neurons. Brain Res. 1976 Sep 17;114(2):293–303. doi: 10.1016/0006-8993(76)90672-7. [DOI] [PubMed] [Google Scholar]
  15. Goldberg R. L., Costa T., Habig W. H., Kohn L. D., Hardegree M. C. Characterization of fragment C and tetanus toxin binding to rat brain membranes. Mol Pharmacol. 1981 Nov;20(3):565–570. [PubMed] [Google Scholar]
  16. Gonatas N. K., Harper C., Mizutani T., Gonatas J. O. Superior sensitivity of conjugates of horseradish peroxidase with wheat germ agglutinin for studies of retrograde axonal transport. J Histochem Cytochem. 1979 Mar;27(3):728–734. doi: 10.1177/27.3.90065. [DOI] [PubMed] [Google Scholar]
  17. Habermann E., Albus U. Interaction between tetanus toxin and rabbit kidney: a comparison with rat brain preparations. J Neurochem. 1986 Apr;46(4):1219–1226. doi: 10.1111/j.1471-4159.1986.tb00641.x. [DOI] [PubMed] [Google Scholar]
  18. Habermann E., Bigalke H., Heller I. Inhibition of synaptosomal choline uptake by tetanus and botulinum A toxin. Partial dissociation of fixation and effect of tetanus toxin. Naunyn Schmiedebergs Arch Pharmacol. 1981 Apr;316(2):135–142. doi: 10.1007/BF00505307. [DOI] [PubMed] [Google Scholar]
  19. Helting T. B., Zwisler O. Structure of tetanus toxin. I. Breakdown of the toxin molecule and discrimination between polypeptide fragments. J Biol Chem. 1977 Jan 10;252(1):187–193. [PubMed] [Google Scholar]
  20. King T. P., Li Y., Kochoumian L. Preparation of protein conjugates via intermolecular disulfide bond formation. Biochemistry. 1978 Apr 18;17(8):1499–1506. doi: 10.1021/bi00601a022. [DOI] [PubMed] [Google Scholar]
  21. Mellanby J., Green J. How does tetanus toxin act? Neuroscience. 1981;6(3):281–300. doi: 10.1016/0306-4522(81)90123-8. [DOI] [PubMed] [Google Scholar]
  22. Nakane P. K., Watanabe T. Distribution of oncodevelopmental markers in neoplastic cells: therapeutic implications. J Histochem Cytochem. 1984 Aug;32(8):894–898. doi: 10.1177/32.8.6747276. [DOI] [PubMed] [Google Scholar]
  23. Pardridge W. M. Recent advances in blood-brain barrier transport. Annu Rev Pharmacol Toxicol. 1988;28:25–39. doi: 10.1146/annurev.pa.28.040188.000325. [DOI] [PubMed] [Google Scholar]
  24. Price D. L., Griffin J., Young A., Peck K., Stocks A. Tetanus toxin: direct evidence for retrograde intraaxonal transport. Science. 1975 May 30;188(4191):945–947. doi: 10.1126/science.49080. [DOI] [PubMed] [Google Scholar]
  25. Rogers T. B., Snyder S. H. High affinity binding of tetanus toxin to mammalian brain membranes. J Biol Chem. 1981 Mar 10;256(5):2402–2407. [PubMed] [Google Scholar]
  26. Schwab M. E., Suda K., Thoenen H. Selective retrograde transsynaptic transfer of a protein, tetanus toxin, subsequent to its retrograde axonal transport. J Cell Biol. 1979 Sep;82(3):798–810. doi: 10.1083/jcb.82.3.798. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Schwab M., Agid Y., Glowinski J., Thoenen H. Retrograde axonal transport of 125I-tetanus toxin as a tool for tracing fiber connections in the central nervous system; connections of the rostral part of the rat neostriatum. Brain Res. 1977 May 6;126(2):211–224. doi: 10.1016/0006-8993(77)90722-3. [DOI] [PubMed] [Google Scholar]
  28. Stoeckel K., Schwab M., Thoenen H. Role of gangliosides in the uptake and retrograde axonal transport of cholera and tetanus toxin as compared to nerve growth factor and wheat germ agglutinin. Brain Res. 1977 Aug 26;132(2):273–285. doi: 10.1016/0006-8993(77)90421-8. [DOI] [PubMed] [Google Scholar]
  29. Stöckel K., Schwab M., Thoenen H. Comparison between the retrograde axonal transport of nerve growth factor and tetanus toxin in motor, sensory and adrenergic neurons. Brain Res. 1975 Nov 28;99(1):1–16. doi: 10.1016/0006-8993(75)90604-6. [DOI] [PubMed] [Google Scholar]
  30. Weller U., Taylor C. F., Habermann E. Quantitative comparison between tetanus toxin, some fragments and toxoid for binding and axonal transport in the rat. Toxicon. 1986;24(11-12):1055–1063. doi: 10.1016/0041-0101(86)90132-7. [DOI] [PubMed] [Google Scholar]
  31. Yavin E., Yavin Z., Kohn L. D. Temperature-mediated interaction of tetanus toxin with cerebral neuron cultures: characterization of a neuraminidase-insensitive toxin-receptor complex. J Neurochem. 1983 May;40(5):1212–1219. doi: 10.1111/j.1471-4159.1983.tb13559.x. [DOI] [PubMed] [Google Scholar]
  32. Zimmerman J. M., Piffaretti J. C. Interaction of tetanus toxin and toxoid with cultured neuroblastoma cells. Analysis by immunofluorescence. Naunyn Schmiedebergs Arch Pharmacol. 1977 Feb;296(3):271–277. doi: 10.1007/BF00498693. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES