84. Plasmid copy number achieved by plasmid-encoded control elements that regulate the initiation of the replication step. For example in stringent plasmid protein Rep A dimerize and binds to origin of replication and donot allow replication more than once.

What mutation may convert this stringent mode of replication in plasmid into relaxed one?

(1) Over expression in repA protein

(2) Mutation in repB dimerization domain

(3) Mutation in repA other than dimerization domain

(4) Gain of function in recognition domain of repA

Background: Stringent vs Relaxed Plasmid Replication Control

Plasmids maintain their copy number in bacterial cells through tightly regulated replication initiation mechanisms.

• Stringent plasmids replicate in coordination with the host chromosome, maintaining a low copy number.

• Relaxed plasmids replicate independently and often achieve high copy numbers.

In stringent plasmids, RepA protein plays a critical role by dimerizing and binding to the origin of replication (ori), inhibiting multiple initiations per cell cycle, thus enforcing low copy number.

Role of RepA Dimerization in Stringent Control

• RepA dimers bind ori DNA, preventing over-initiation of replication.

• This negative feedback ensures replication occurs only once per cell cycle per plasmid copy.

• Dimerization is essential for this inhibitory function.

Effect of Mutations on Replication Control

• Mutation in the RepA dimerization domain disrupts the ability of RepA to form dimers.

• Without dimerization, RepA cannot effectively bind ori to inhibit replication initiation.

• This loss of negative regulation converts the plasmid from a stringent to a relaxed replication mode, allowing multiple rounds of replication and increasing plasmid copy number.

Why Other Mutations Are Less Likely to Cause Relaxed Replication

• Overexpression of RepA protein would likely enhance repression, maintaining or strengthening stringent control.

• Mutation in RepA outside the dimerization domain may not affect dimer formation or ori binding significantly.

• Gain of function in RepA recognition domain would likely increase binding and repression, not reduce it.

Summary Table

Conclusion

The mutation that most likely converts stringent plasmid replication into relaxed replication is a mutation in the RepA dimerization domain. This mutation prevents RepA from forming dimers that inhibit replication initiation, leading to increased plasmid copy number and relaxed control.

Keywords

plasmid replication, stringent control, relaxed control, RepA protein, dimerization domain, plasmid copy number, replication initiation, bacterial plasmids, mutation effects

Correct answer:
(2) Mutation in repB dimerization domain