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J. Biochem, 2004, Vol. 135, No. 4 509-516
© 2004 The Japanese Biochemical Society


BIOCHEMISTRY

Characterization and cDNA Cloning of Monomeric Lectins That Correspond to the B-Chain of a Type 2 Ribosome-Inactivating Protein from the Bark of Japanese Elderberry (Sambucus sieboldiana)

Maria Angeles Rojo*,1, Hanae Kaku1, Naoko Ishii-Minami1, Eiichi Minami1, Misa Yato1,2, Shigeru Hisajima2, Takeshi Yamaguchi§,1 and Naoto Shibuya,3

1 Department of Biochemistry, National Institute of Agrobiological Resources, and 2 Institute of Applied Biochemistry, University of Tsukuba, Tsukuba, Ibaraki 305; and 3 Department of Life Sciences, School of Agriculture, Meiji University, 1-1-1 Higashi-mita, Tama-ku, Kawasaki, Kanagawa 214-8571

Two monomeric lectins, SSA-b-3 and SSA-b-4, were purified from the bark tissue of Japanese elderberry, Sambucus sieboldiana. SDS-PAGE of the purified lectins showed the presence of single bands of 35 and 33 kDa for SSA-b-3 and SSA-b-4, respectively, irrespective of the presence of reducing agent. MS analysis as well as gel filtration of these lectins indicated that they exist mostly as monomeric lectins. Analysis of the N-terminal amino acid sequences of SSA-b-3 and SSA-b-4 yielded an identical sequence, indicating their close structural relationship. Four cDNA clones with extensive homology were obtained from the bark cDNA library and indicated to encode SSA-b-3 or SSA-b-4 from the comparison with the N-terminal sequences of these lectins. These clones were classified into two groups, three for SSA-b-3 and one for SSA-b-4, based on the predicted isoelectric points. The amino acid sequences of the encoded polypeptides were almost identical with the B-chain of a type 2 ribosome-inactivating protein from the same bark tissue, sieboldin-b, except for the absence of a small peptide containing a cystein residue, which is critical for the heteromeric dimerization with an A-subunit. Carbohydrate binding specificity and biological activity of these lectins are also reported.

* Present address: Universidad de Valladolid, E-47005 Valladolid, Spain.

§ Present address: National Agricultural Research Center, Joetsu, Niigata 943-0193.

To whom correspondence should be addressed. Tel/Fax: +81-44-934-7039, E-mail: shibuya{at}isc.meiji,ac.jp


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