Thermostable peroxidase activity with a recombinant antibody L chain‐porphyrin Fe(III) complex

FEBS Letters

Harada, Akira²  Takagi, Masahiro¹  Kamachi, Mikiharu²  Kohda, Katsunori¹  Imanaka, Tadayuki¹  Yamaguchi, Hiroyasu²  Hamuro, Takuya¹ 

[1] Department of Biotechnology, Faculty of Engineering, Osaka University, 2-1 Yamada-oka, Suita, Osaka 565, Japan;Department of Polymer Science, Faculty of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka 560, Japan

关键词: Catalytic antibody;    Peroxidase;    Porphyrin;    L chain;   

DOI  :  10.1016/0014-5793(95)01224-3

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

In order to engineer a new type of catalytic antibody, we attempt to use a monoclonal antibody L chain as a host protein for a porphyrin. TCPP (meso-tetrakis(4-carboxyphenyl)porphyine) was chemically synthesized and Balb/c mice were immunized using TCPP as a hapten. Two hybridoma cells (03-1, 13-1), that produce monoclonal antibody against TCPP, were obtained. Genes for both H and L chains of monoclonal antibodies were cloned, sequenced and overexpressed using E. coli as a host. ELISA and fluorescence quenching method show that the independent antibody L chains from both Mab03-1 and Mab13-1 have specific interaction with TCPP. Furthermore, the recombinant antibody L chain from Mab13-1 exhibits much higher peroxidase activity than TCPP Fe(III) alone. The enzyme activity was detectable with pyrogallol and ABTS (2,2-azinobis-3-ethylbenzthiazolin-6-sulfonic acid) but not with catechol. This new catalytic antibody was extremely thermostable. Optimum temperature of the peroxidase reaction by the complex of 13-1L chain and TCPP Fe(III) was 90°C, while that the TCPP Fe(III) alone was 60°C.

【 授权许可】

Unknown