Production of a bifunctional hybrid molecule B72.3/metallothionein-1 by protein engineering.
A hybrid anti-tumour B72.3 antibody/metallothionein protein B72.3MT-1 was produced by the construction of the expression vector mpSV2neo-EP1-B72.3MT-1. This vector contained the neo gene as a selection marker, the murine immunoglobulin promoter and enhancer, and the hybrid B72.3 heavy chain gene fragment with mouse metallothionein-1 cDNA gene ligated into its CH2 domain. The expression vector was transfected to the heavy chain loss mutant B72.3Mut(K) cell line. The hybrid protein B72.3MT-1 was purified from transfectant supernates using a Protein G column. We showed that the hybrid protein retained the binding reactivity for the TAG72 antigen as the original B72.3 antibody, and the metal-binding capacity of the native metallothionein molecule. Therefore, the bifunctional hybrid protein B72.3MT-1 may be very useful in cancer imaging when labelled with radionuclides such as 99mTc.
Xiang
J
J
Saskatoon Cancer Center, Department of Microbiology, University of Saskatchewan, Canada.
Koropatnick
J
J
Qi
Y
Y
Luo
X
X
Moyana
T
T
Li
K
K
Chen
Y
Y
D000230:Adenocarcinoma / Q000276:immunology; D000912:Antibodies, Neoplasm / Q000096:biosynthesis*; D000918:Antibody Specificity; D001483:Base Sequence; D002460:Cell Line; D003110:Colonic Neoplasms / Q000276:immunology; D004247:DNA / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D005822:Genetic Vectors; D006801:Humans; D007124:Immunoenzyme Techniques; D008668:Metallothionein / Q000235:genetics; D008969:Molecular Sequence Data; D015202:Protein Engineering / Q000379:methods*; D055503:Protein Multimerization
D000912:Antibodies, Neoplasm; D004247:DNA; D008668:Metallothionein