Question

A mutation in the operator upstream of the lac promoter (O₃) reduces the repression of the lac promoter by the lac repressor by about 3-fold. Why is this? Describe an intergenic suppressor of such an O₃ mutation.

Answer 1

The lac operon has 3 lac repressor recognition sites in a stretch of 500bp. They are at the positions of three operator sites , O1, O2 and O3. O3 lies with in the lac I gene,which is 93 bp, upstream of O1 which lies in the promoter. O2 lies 401 bp upstream of O1 within the lac z gene.

The effect of systematic destruction of lac operator of the chromosomal lac operon by reducing the repression of the lac operon by lac repressor. Absence of pseudo-operator O3 decreases repression by wild type tetrameric lac repressor approximately 3 fold . Absence of pseudo-operator decreases repression greater than 50 fold. This indicates that cooperative interaction between lac operators is due to DNA loop formation mediated by tetrameric lac repressor.

Intergenic suppression relieves the effects of a mutation in one gene by a mutation somewhere else within the genome. This mutation involves a a change in protein structure caused by first mutation. An intergenic suppressor mutation occurs in different genes from where the first mutation occurs. This involves a change in the expression of one gene that compensates other gene product.

Additional constructs in which the auxiliary lac operator O3 replaced by 7-2-7 heptamer pairs. The lac repressor consists of two DNA binding dimers assembled as a homotetramer. The lac operon control region might provide means to study specific interactions between phospho-Narl dimers. Accordingly, site -slecific mutagenesis to replace the auxiliary operator O3-lac with 7-2-7 heptamer pairs from the nirab or napF control regions. This operators were placed in the various combinations of O3 and O1 substituted lac control regions.