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For the second step of trypanothione synthesis glutathionylspermidine is bound in a way that preferentially allows N1-glutathionylation of N8-glutathionylspermidine, classifying N8-glutathionylspermidine as the favoured substrate

1 Step 1: Ethanol Acetaldehyde Alcohol dehydrogenase (ADH) converts ethanol to acetaldehyde, a far more toxic molecule