Ruf 5 - Main Points Lecture 4 Lac repressor biochemistry...

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Lac repressor biochemistry: functions as a tetramer, each dimer binds an operator site, each operator site consists of 2 half-sites each of which is bound by a monomer of the repressor Binding of inducer alters protein confirmation: prevents cooperative binding to half-sites, reduces affinity of protein for operator site DNA-protein binding assays: footprinting and EMSA Differential protein levels of components of operon: translational polarity and rate of RNA degradation Main Points: Lecture 4
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Genotype B-gal made Inducible or Constitutive? Grow on Lac only? i- p+ O+ Z+ Y+ i+ p+ Oc Z- Y+ 1. Can B gal be made? a. Do you have a wild-type Z present ? Yes b. Do you have a functional promoter ? Yes 1. Ind or Con (refers only to B gal not permease)? a. Do you have a functional repressor (i+) ? Yes b. Do you have a functional operator ? Yes 1. Grow on Lac only? a. Can you make both Z and Y ? Yes
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Genotype B-gal made Inducible or Constitutive? Grow on Lac only? i- p+ O+ Z+ Y+ i+ p+ Oc Z- Y+ i+ p- O+ Z+ Y+ i- p+ O+ Z- Y+ i+ Oc Z- Y+ / i+ Z+ Y- Yes Ind Yes i- p- Oc Z+ Y+/ i- Z+ Y- Yes Con NO 1. Can B gal be made? a. Do you have a wild-type Z present ?
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