Recombinant botulinum neurotoxin A heavy chain-based delivery vehicles for neuronal cell targeting.
The long half-life of botulinum neurotoxin serotype A (BoNT/A) in cells poses a challenge in developing post-exposure therapeutics complementary to existing antitoxin strategies. Delivery vehicles consisting of the toxin heavy chain (HC), including the receptor-binding domain and translocation domain, connected to an inhibitory cargo offer a possible solution for rescuing intoxicated neurons in victims paralyzed from botulism. Here, we report the expression and purification of soluble recombinant prototype green fluorescent protein (GFP) cargo proteins fused to the entire BoNT/A-HC (residues 544-1295) in Escherichia coli with up to a 40 amino acid linker inserted between the cargo and BoNT/A-HC vehicle. We show that these GFP-HC fusion proteins are functionally active and readily taken up by cultured neuronal cells as well as by neuronal cells in mouse motor nerve endings.
Department of Microbiology, University of Illinois at Urbana-Champaign, Champaign, IL, USA.
Mengfei
M
Ho
Li-Hsin
LH
Chang
Melissa
M
Pires-Alves
Baskaran
B
Thyagarajan
Jordan E
JE
Bloom
Zhengrong
Z
Gu
Karla K
KK
Aberle
Sasha A
SA
Teymorian
Yuka
Y
Bannai
Steven C
SC
Johnson
Joseph J
JJ
McArdle
Brenda A
BA
Wilson
D000818:Animals; D019274:Botulinum Toxins, Type A / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D004337:Drug Carriers / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D016503:Drug Delivery Systems / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics; D051379:Mice; D009046:Motor Neurons / Q000166:cytology / Q000378:metabolism; D009469:Neuromuscular Junction / Q000166:cytology; D009474:Neurons / Q000166:cytology* / Q000378:metabolism*; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D012018:Reflex / Q000187:drug effects; D014034:Toes / Q000502:physiology
D004337:Drug Carriers; D011993:Recombinant Fusion Proteins; D019274:Botulinum Toxins, Type A