Localization of two domains of a mutant form of Escherichia coli protein L7/L12 that binds the large ribosomal subunit as a single dimer.

Escherichia coli ribosomal protein L7/L12 occurs on the large subunit as two dimers: one dimer is extended and comprises the stalk, while the second dimer is folded and occupies a site on the subunit body. A variant protein, in which all 18 amino acids of the flexible hinge region that links separate N-terminal and C-terminal domains of L7/L12 has been deleted, binds the subunit as a single dimer and does not generate stalks that are visible in electron micrographs. Monoclonal antibodies directed against each domain of the protein have been used to localize the variant in electron micrographs of 50S subunits. Both C-terminal domains are seen at a shoulder of the subunit, near its edge as viewed in the most common quasisymmetric projection. N-terminal domains are placed on the subunit body, about 50 A from the C-terminal domains. The antibody to the N-terminal domain also causes dissociation of the variant dimer from the particle and the formation of oligomeric antibody-protein dimer complexes. Similar complexes were seen previously (Olson HM et al (1986) J Biol Chem 261, 6924-6936) when this antibody induced dissociation of one dimer of the native protein. We conclude that the shortened variant most probably occupies the lower-affinity site on the subunit that is normally filled by the stalk dimer.

Evidence Against Amplification of Four Genes in Giant Cells Induced by Meloidogyne incognita.

Giant-cell DNA was isolated from pea (Pisum sativum) inoculated with Meloidogyne incognita and used in slot blots to test for selective sequence amplification. Four sequences representing low (ribulose 1,5-bisphosphate carboxylase and actin), mid-level (histone 3), and highly repetitive (large ribosomal repeat) sequence DNA were used as probes. Known amounts of root-tip DNA and giant-cell DNA were blotted onto hybridization membranes and probed. The signal strength on autoradiographs containing equal amounts of root-tip DNA and giant-cell DNA were compared with a scanning densitometer. No difference in signal strength between equal amounts of root-tip DNA and giant-cell DNA was found. Thus, for the probes tested, there is no difference in copy number and, hence, no selective DNA sequence amplification has occurred.