Hegde R, Podder S K
Department of Biochemistry, Indian Institute of Science, Bangalore.
Eur J Biochem. 1998 Jun 15;254(3):596-601. doi: 10.1046/j.1432-1327.1998.2540596.x.
Seeds of Ricinus communis contain two types of lectins; the toxin ricin (approximately 60 kDa) and R. communis agglutinin (approximately 120 kDa). The toxin is a heterodimer composed of a toxic A subunit and a lectin B subunit, while R. communis agglutinin is a tetramer, constituted of two ricin-like dimers held together by non-covalent forces. The lactamyl Sepharose affinity-purified ricin consists of two major groups of variants designated ricin II and III [Hegde, R. & Podder, S. K. (1992) Eur. J. Biochem. 204, 155-164]. The purified A subunits of all the variants of ricins and R. communis agglutinin show heterogeneity in the molecular mass as shown for ricin before [Fulton, J. R., Blakey, C. D., Knowles, P. P., Uhr, J. W., Thorpe, P. E. & Vitetta, E. S. (1986) J. Biol. Chem. 261, 5314-5319]. Since the isoelectric points of the R. communis agglutinin variants fall between the isoelectric points of ricin II and III, we investigated the possibility that this lectin is made up of ricin II and III. The isoelectric points of the purified B subunits of R. communis agglutinin matched well with those of ricin II and III on urea-polyacrylamide isoelectric focussing gel. Further, two-dimensional electrophoretic analysis of the ricin constituants of R. communis agglutinin in the presence of 9 M urea, showed two protein bands, differing by nearly pH 2 in their isoelectric points, the more alkaline one corresponding to that of ricin III analyzed under the same conditions, while the other, although a higher molecular mass variant, corresponding well with ricin II in its isoelectric point. Based on these results and those obtained from adenine binding to A chains of both ricin and R. communis agglutinin, we provide a plausible evolutionary relationship between R. communis agglutinin and two groups of ricin variants; ricin II and III. The model predicts that one half of R. communis agglutinin is derived from ricin I and II, and the other half from ricin III. The results presented, contrary to the existing notion, unequivocally show that the two halves of R. communis agglutinin are not identical protein units, but differ both in surface charge and molecular mass.
毒素蓖麻毒蛋白（约60 kDa）和蓖麻凝集素（约120 kDa）。毒素是一种异二聚体，由有毒的A亚基和凝集素B亚基组成，而蓖麻凝集素是一种四聚体，由两个蓖麻毒蛋白样二聚体通过非共价力结合在一起构成。内酰胺琼脂糖亲和纯化的蓖麻毒蛋白由两组主要的变体组成，分别命名为蓖麻毒蛋白II和III [Hegde, R. & Podder, S. K. (1992) Eur. J. Biochem. 204, 155 - 164]。所有蓖麻毒蛋白变体和蓖麻凝集素的纯化A亚基在分子量上都表现出异质性，如之前对蓖麻毒蛋白的研究所示 [Fulton, J. R., Blakey, C. D., Knowles, P. P., Uhr, J. W., Thorpe, P. E. & Vitetta, E. S. (1986) J. Biol. Chem. 261, 5314 - 5319]。由于蓖麻凝集素变体的等电点落在蓖麻毒蛋白II和III的等电点之间，我们研究了这种凝集素由蓖麻毒蛋白II和III组成的可能性。在尿素 - 聚丙烯酰胺等电聚焦凝胶上，蓖麻凝集素纯化B亚基的等电点与蓖麻毒蛋白II和III的等电点匹配良好。此外，在9 M尿素存在下对蓖麻凝集素的蓖麻毒蛋白成分进行二维电泳分析，显示出两条蛋白带，其等电点相差近2个pH单位，碱性较强的一条与在相同条件下分析的蓖麻毒蛋白III相对应，而另一条虽然是分子量较高的变体，但其等电点与蓖麻毒蛋白II非常吻合。基于这些结果以及从腺嘌呤与蓖麻毒蛋白和蓖麻凝集素的A链结合获得的结果，我们提出了蓖麻凝集素与两组蓖麻毒蛋白变体（蓖麻毒蛋白II和III）之间合理的进化关系。该模型预测，蓖麻凝集素的一半源自蓖麻毒蛋白I和II，另一半源自蓖麻毒蛋白III。所呈现的结果与现有观点相反，明确表明蓖麻凝集素的两半不是相同的蛋白质单元，而是在表面电荷和分子量上都有所不同。
