Our research program is aimed at elucidating the mechanism of the steps involved in the transport of macromolecular DNA through the cell envelope of Haemophilus influenzae in the course of genetic transformation. The proposed experiments are aimed at providing direct evidence for the participation of cellular proteins in the process of DNA transport. The proteins interacting with transforming DNA will be isolated by use of specific affinity techniques. Photodynamic crosslinking of bound proteins with transforming DNA will be exploited to further characterize the interacting species. Antibodies directed against the purified DNA binding proteins will be used to probe the action of these proteins in situ. A periplasmic DNA binding protein will be characterized and studied by these techniques. Mutants conditionally defective in the transformation process will be isolated using a rapid screening technique developed during this project. Membrane vesicles active in the uptake of DNA will be employed as antigens and studied by both immunological and affinity methods, and the extent to which these vesicles behave like the intact cell will be studied.