Harvard College

University of Pennsylvania

Feifan Yu

Min Dong

Andrew Tsourkas

Burcin Altun

Hejia Henry Wang

The invention relates to proximity-based sortase-mediated protein purification and ligation. Specifically, the invention relates to techniques that links protein expression/purification with conjugation to therapeutic agents, imaging agents, or linkers.

Proximity-based sortase-mediated protein purification and ligation

Disclosed herein are means for the detection and characterization of neurotoxins such asneurotoxin (BoNT) or tetanus neurotoxin. The present disclosure provides methods for determining potency and activity of neurotoxins in vitro and in vivo. Also disclosed are polypeptides comprising N- and C-terminal fragments of a reporter protein that are split by a linker comprising a neurotoxin cleavage site. Cleavage of the linker by a neurotoxin decreases reporter protein activity, thereby indicating activity of the neurotoxin. Compositions and kits comprising the disclosed polypeptides, nucleic acids comprising nucleotide sequences encoding such polypeptides, and cells expressing such polypeptides are also disclosed.

In vitro and cell based assays for measuring the activity of botulinum neurotoxins

Disclosed herein are means for the detection and characterization of neurotoxins such as botulinum neurotoxin (BoNT) or tetanus neurotoxin. The present disclosure provides methods for determining potency and activity of neurotoxins in vitro and in vivo. Also disclosed are polypeptides comprising N- and C-terminal fragments of a reporter protein that are split by a linker comprising a neurotoxin cleavage site. Cleavage of the linker by a neurotoxin decreases reporter protein activity, thereby indicating activity of the neurotoxin. Compositions and kits comprising the disclosed polypeptides, nucleic acids comprising nucleotide sequences encoding such polypeptides, and cells expressing such polypeptides are also disclosed.

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Aberdeen, NJ, United States of America

Feifan Yu