Sialic acid binding lectin from Rana catesbeiana oocyte

(cSBL) is known to have anti-tumor activity. cSBL is a multifunctional protein and is recognized as a pyrimidine basespecific ribonuclease. In respect to its ribonuclease activity, cSBL belongs to the RNase A superfamily, and is a very heat-stable enzyme. It retains 100% activity after heating at 100 °C for 5 min. Four RNases have been isolated from the Rana family of frogs, and their primary structures have been determined to date. All have a very similar primary structure, as shown in Fig. 1. Each of these RNases has antitumor activity, except the one from bullfrog liver. Among them, the RNase from R. pipiens (onconase) has been studied most extensively and is in phase III clinical trials as an antitumor drug. Nitta et al. reported on the anti-tumor activity of cSBL. The mechanism of the anti-tumor action of frog oocyte RNases was studied using cSBL and onconase, but has not yet been fully elucidated. Nitta et al. proposed that cSBL interacts with a sialoglycoprotein on the surface of tumor cells, is then internalized and works in the cells. The proposal was based on the fact that cSBL was not effective against murine leukemia P388 cells treated with sialidase. On the other hand, Wu, N. Y. reported (cited in ref. 11) that the anti-tumor activity of onconase is not affected by pretreatment with sialidase, in spite of its structural similarity with cSBL. Nitta et al. reported that the chemical modification of cSBL with a water soluble carbodiimide (EDC) in the presence of glycine methylester increased its anti-tumor action. In the previous paper, we reported that (i) modification of the carboxyl groups of cSBL with EDC in the presence of nucleophiles enhanced the cytotoxic activity toward P388 cells. The enhancing effect is dependent on the increase in positive net charge, thus the cytotoxicity was greatest in the order of cSBL modified in the presence of ethylenediamine (EDC-ED cSBL).glycine methylester (EDC-GM cSBL). taurine (EDC-TA cSBL). (ii) cSBL with a higher net charge is more efficiently internalized into the P388 cells than the native cSBL. However, we have not been able to define which carboxyl group is chemically modified in each sample. (iii) EDC-ED cSBL hydrolyzed RNA hybridized with RNA or DNA more effectively than the native cSBL. (iv) The base specificity especially that of the B2 site of EDC-ED cSBL, was altered by the chemical modification.