Reductive cleavage of tetanus toxin and botulinum neurotoxin A by the thioredoxin system from brain. Evidence for two redox isomers of tetanus toxin.
Inhibition of neurotransmitter release by tetanus toxin and botulinum neurotoxin A can be mimicked by intracellular application of the corresponding toxin light chains. The aim of this study was to determine whether the two-chain toxins are reduced by brain preparations to yield free light chains which would represent the ultimate toxins. The interchain disulfide of two-chain tetanus toxin was cleaved by rat cortex homogenate fortified with NADPH. Reduction was promoted further by addition of thioredoxin. Thioredoxin reductase was demonstrated in and purified from porcine brain cortex. The thioredoxin system which consisted of purified enzyme, thioredoxin and NADPH reduced both toxins. The resulting light chains appeared homogeneous in SDS gel electrophoresis. The complementary heavy chain of tetanus but not of botulinum toxin migrated in two bands, the faster one with the velocity of heavy chain obtained by chemical reduction. The major, slower form was converted into the faster by chemical but not by enzymatic reduction. Tetanus toxin, whether in its single-chain or two-chain version also occurred in two forms which differed by their electrophoretic mobility. The two forms of single-chain toxin were interconverted by chemical reduction or oxidation but not by the thioredoxin system. It is concluded that a) a thioredoxin system in brain tissue reduces the interchain disulfide of two-chain tetanus toxin and botulinum neurotoxin A, b) tetanus toxin but not botulinum neurotoxin A consists of two electrophoretically distinct forms which differ by the thiol-disulfide status of their heavy chains, c) the disulfide loop within the heavy chain of tetanus toxin is resistant to the thioredoxin system.
Kistner
A
A
Rudolf-Buchheim-Institut für Pharmakologie, Justus-Liebig-Universität, Giessen, Federal Republic of Germany.
Habermann
E
E
D000818:Animals; D001905:Botulinum Toxins / Q000737:chemistry / Q000378:metabolism*; D001921:Brain / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D007535:Isomerases / Q000378:metabolism; D007536:Isomerism; D010084:Oxidation-Reduction; D019704:Protein Disulfide-Isomerases; D051381:Rats; D013552:Swine; D013744:Tetanus Toxin / Q000737:chemistry / Q000378:metabolism*; D013880:Thioredoxin-Disulfide Reductase / Q000302:isolation & purification / Q000378:metabolism*; D013879:Thioredoxins / Q000378:metabolism*
D013744:Tetanus Toxin; D013879:Thioredoxins; D013880:Thioredoxin-Disulfide Reductase; D001905:Botulinum Toxins; D007535:Isomerases; D019704:Protein Disulfide-Isomerases