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J. Biochem, 1974, Vol. 76, No. 6 1227-1234
© 1974 Japanese Biochemical Society

research-article

Studies on Rat Liver Catalase

VIII. Isolation and Translation of Catalase Messenger RNA*

Terufumi SAKAMOTO and Tokuhiko HIGASHI

Department of Biochemistry, School of Pharmaceutical Sciences, Showa University Shinagawa-ku, Tokyo 142

Messenger RNA directing the biosynthesis of catalase [EC 1.11.1.6 [EC] ] has been purified from rat liver. The procedure consists of: (1) isolation of catalase-synthesizing polysomes by immunoprecipitation, (2) extraction of RNA from the polysomes with phenol, and (3) adsorption of mRNA on nitrocellulose filters.

Isolated catalase mRNA was translated by a cell-free system using ribosomes prepared from eggs of a brine shrimp, Artemia salina, and cell sap of rat liver. In this system the mRNA directed the incorporation of 14C-leucine into catalase at least 70% as efficiently as into total proteins.

Catalase mRNA was obtained from both free and membrane-bound polysomes. The main component in the purified catalase mRNA preparation was pulse-labeled with [3H]orotic acid, and was assumed to be about 15S on the basis of sucrose gradient centrifugation. Nitrocellulose filters were effective in the purification of catalase mRNA, suggesting that this messenger contains a poly (A) segment, as is the case in some other mammalian messengers.

*This work was supported in part by a Grant-in-Aid for Scientific from the Ministry of Education, for which the authors are greatly indebted.


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