Patent ID: 8454976

Claim:
A method of making an activated di-chain polypeptide, said method comprising: (a) providing a single-chain fusion protein comprising (i) a clostridial neurotoxin light chain protease, wherein said light chain protease includes the active proteolytic enzyme domain of the light chain and is capable of cleaving one or more vesicle or plasma-membrane associated proteins essential for exocytosis; (ii) a clostridial neurotoxin heavy chain H N portion, wherein said H N portion is capable of translocating (i) into a mammalian cell; and (iii) a protease cleavage site that has been introduced into the single-chain fusion protein; (b) contacting the single-chain fusion protein with a protease that cleaves said single-chain fusion protein at the protease cleavage site; and (c) cleaving said single-chain fusion protein with said protease, thereby providing an activated di-chain polypeptide wherein (i) and (ii) are linked together by a disulphide bond.