Engineering of a mini-trichosanthin that has lower antigenicity by deleting its C-terminal amino acid residues.
Trichosanthin is a ribosome-inactivating protein that possesses antitumor and antiviral activities. Clinical trials of trichosanthin on AIDS patients, however, elicit anaphylactic reactions. To reduce the antigenicity of trichosanthin as a drug while preserving its biological activity, the C-terminal domain (residues 203 to 247), which contains a putative antigenic site, was systemically deleted. We have found that the minimum length of trichosanthin that can fold into an active conformation is residue 1 to 240. The mini-trichosanthin (C7) generated by deleting the last seven C-terminal amino acid residues has 2.7-fold decrease in antigenicity, 10-fold reduction in in vitro ribosome-inactivation activity, and in vivo cytotoxicity toward K562 cells, and 2-fold reduction in abortificient activity. Structural analyses of C7 indicate decrease in the helix content, increased exposure of Trp192, and lower thermodynamic stability. The deletion of the C-terminal residues (Leu241 to Ala247) probably perturbs local structure of the C-terminal antigenic epitope that results in the decrease in antigenicity and activities of C7.
Chan
S H
SH
Department of Biochemistry, Department of Physiology, Chinese University of Hong Kong, Shatin, N.T., Hong Kong, China.
Shaw
P C
PC
Mulot
S F
SF
Xu
L H
LH
Chan
W L
WL
Tam
S C
SC
Wong
K B
KB
D000020:Abortifacient Agents, Nonsteroidal / Q000276:immunology*; D000595:Amino Acid Sequence; D019380:Anti-HIV Agents / Q000276:immunology*; D000972:Antineoplastic Agents, Phytogenic / Q000276:immunology*; D002942:Circular Dichroism; D019791:Guanidine / Q000494:pharmacology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011489:Protein Denaturation; D015202:Protein Engineering; D017433:Protein Structure, Secondary; D012270:Ribosomes / Q000187:drug effects; D017384:Sequence Deletion; D015978:Trichosanthin / Q000235:genetics / Q000276:immunology*
D000020:Abortifacient Agents, Nonsteroidal; D019380:Anti-HIV Agents; D000972:Antineoplastic Agents, Phytogenic; D015978:Trichosanthin; D019791:Guanidine