Disulphide Bonding in a Stylar Self‐Incompatibility Ribonuclease of Nicotiana Alata

Abstract
Many flowering plants have developed a self‐incompatibility mechanism, which is controlled by a single polyallelic locus (the S‐locus), to prevent inbreeding. The products of the S‐locus in the styles of solanaceous plants are an allelic series of glycoproteins with RNase activity [McClure, B. A., Haring, V., Ebert, P. R., Anderson, M. A., Simpson, R. J., Sakiyama, F. & Clarke, A. E. (1989) Nature 342, 955–957]. These S‐RNases show some amino‐acid‐sequence similarity with two fungal RNases (T2 and Rh), including the presence of two active‐site His residues, which suggests a common three‐dimensional structure. Disulphide bonding is important in the maintenance of the three‐dimensional structure of the fungal RNases [Kurihara, H., Mitsui, Y., Ohgi, K., Irie, M., Mizuno, H. & Nakamura, T. (1992) FEBS Lett. 306, 189–192] and the S‐RNases [Tsai, D. S., Lee, H.‐S., Post, L. C., Kreiling, K. M. & Kao, T.‐H. (1992) Sex. Plant Reprod. 5, 256–263]. We have used the S2‐allele RNase of Nicotiana alata, which has nine Cys residues, to establish the pattern of disulphide bonding. The disulphide bonds Cys16–Cys21, Cys45–Cys94, Cys153–Cys182 and Cys165–Cys176 are consistent with the S2‐RNase having a similar three‐dimensional structure to RNase Rh. A free Cys residue (Cys95) adjacent to Cys45‐Cys94 promotes a rapid specific disulphide migration when the protein is exposed to denaturing conditions.