KINETICS OF CONJUGATIVE TRANSFER - A STUDY OF THE PLASMID PXO16 FROM BACILLUS-THURINGIENSIS SUBSP ISRAELENSIS

The aggregation-mediated conjugation system of Bacillus thuringiensis subsp. israelensis, encoded by the 200-kb plasmid pXO16, is highly pot ent in transferring itself and efficient in mobilizing other nonconjug ative plasmids. The present study reveals some salient features of thi s conjugation system. Our observations can be summarized as follows: ( i) The conjugative transfer takes about 3 1/2 to 4 min. For a 200-kb p lasmid this corresponds to about 1 kb per second. (ii) The ability to transfer the plasmid seems to be evenly distributed among the donors. (iii) Functionally, the mating complex was found to consist of one don or and one recipient cell, even though aggregates comprising thousands of interconnected cells are formed. (iv) Having donated the plasmid, the donor needs a ''period of recovery'' of about 10 min before it can redonate the plasmid. (v) Secondary transfer, i.e., transfer from new ly formed transconjugants, is delayed about 40 min. This maturation ti me exceeds the generation time, and it may indicate that to display do nor activity, a surface protein (the aggregation substance) has to be uniformly incorporated into the cell wall. Lastly, we found that when the experiments were sufficiently short and when the recipient cells w ere in excess compared with the donors, the process of conjugation cou ld be reasonably described by a kinetic model analogous to the Michael is-Menten model for enzyme catalysis. This allowed us to estimate (vi) the maximal conjugation rate to be about 0.05 transconjugant per dono r per minute, and (vii) the K-m value, i.e., the concentration of reci pient that results in half of the maximal conjugation rate, to be abou t 4 x 10(6) recipients/ml. (C) 1998 Academic Press.