Kinetics of conjugative transfer: a study of the plasmid pXO16 from Bacillus thuringiensis subsp. israelensis

Abstract

The aggregation-mediated conjugation system of Bacillus thuringiensis subsp. israelensis, encoded by the 200-kb plasmid pXO16, is highly potent in transferring itself and efficient in mobilizing other nonconjugative plasmids. The present study reveals some salient features of this 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 plasmid 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 donor 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 newly formed transconjugants, is delayed about 40 min. This maturation time exceeds the generation time, and it may indicate that to display donor 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 were in excess compared with the donors, the process of conjugation could be reasonably described by a kinetic model analogous to the Michaelis-Menten model for enzyme catalysis. This allowed us to estimate (vi) the maximal conjugation rate to be about 0.05 transconjugant per donor per minute, and (vii) the Km value, i.e., the concentration of recipient that results in half of the maximal conjugation rate, to be about 4 x 10(6) recipients/ml.