notes 18

Molecular Biology Chapter 18 Control of Gene Expression in Bacteria Gene Expression Some genes are constitutively expressed Which genes might fall into this category? All genes are not expressed all of the time Cells can control if a gene is expressed Cells can control how much it is expressed Gene regulation in Arabidopis thaliana Some genes regulate other genes About 27,000 genes total About 1,200 regulatory Why are genes regulated? Cell differentiation All cells have the same genome, do all cells need the same genes expressed? Genes can be expressed only during certain stages of development Why have different versions of hemoglobin? Gene expression can change due to environmental variation DNA mRNA Protein Control can occur between any of these steps (Arrow 1) Transcriptional control: RNA polymerase is prevented from binding to the promoter Why would this be a good place for regulation? (Arrow 2) Translational control: mRNA is degraded at a different rate or translation is not initiated Why would this be a good place for regulation? (Protein) Post-Translational control: protein is not properly folded to its functional conformation Why would this be a good place for regulation? Ecology of E.coli Successful growth and reproduction requires efficient use of energy Also good to exploit a wide variety of energy sources Exploiting an energy source isn’t free Transport proteins, enzymes Different energy sources need different metabolic machinery Different enzymes for glucose, lactose, sucrose, etc… Glucose is preferred Lactose metabolism Disaccharide made of glucose and galactose E.coli uses ?-galactosidase to break the lactose down into glucose and galactose Do you think ?-galactosidase is a constitutive gene? Experiment: Grow E.coli on media with glucose or media with lactose Which is more likely to make ?-galactosidase? Identifying mutants in E.coli To study the proteins involved in lactose metabolism, identify mutants that can’t use lactose Make ‘em yourself! (Monod) Also used indicator plates Colonies with ?-galactosidase turn yellow Colonies with ?-galactosidase deficiency are white 3 mutations in lactose metabolism Cells cant break apart lactose into glucose and galactose Gene= lacZ, mutant= lacZ Cells don’t import lactose Gene= lacY, mutant= lacY, protein= galactoside permease Cells express the lacZ and lacY genes all the time, even when lactose isn’t present Gene= lacI, mutant= lacI Constitutive mutant 2 mutations in functional genes 1 mutation in a regulatory gene Functiona; genes (lacZ and lacY) are linked together in on operon Operon= group of genes under the transcription control of 1 promoter mRNA is polycistronic, includes code for multiple proteins lac operon RNA polymerase attaches to the lac operon promoter, mRNA is made for lacZ, lacY, and lacA as a single molecule lacA= transacetylase, exports sugars when they are too abundant Why express these 3 genes together? lacI is controlled by its own promoter Protein= repressor Repressor binds to operator Operator is between the promoter and the lacZ, lacY, and lacA genes Physically stops RNA polymerase from transcribing the lac genes Negative control: regulatory protein stops transcription Do you think lacI is a constitutively expressed gene? Need to regulate the behavior of the repressor protein What would be a good signal to tell the repressor to let go of the operator? Repressor protein has 4 subunits Each subunit can bind a lactose molecule Allosteric regulation The new conformation doesn’t fit operator anymore, repressor lets go RNA polymerase is free to transcribe the lac genes Lactose = inducer Enzymes that break down lactose are made Lactose concentration in the cell drops Less lactose to bind to repressor returns to its original form, binds to operator Key points about negative control Physical interaction between repressor and the RNA polymerase Changes in the level of negative control happen post-translation Repressor is always made, its structure is altered Allows for rapid change in the expression of the lac genes lacI and repressor aren’t the only mechanism for control Upstream of the lac promoter is the CAP binding site CAP= catabolite activator protein Transcription is enhanced when CAP binds to the CAP binding site Lac promoter is weak Promoters may not cause RNA polymerase to transcribe at the maximum possible rate CAP exerts positive control; causes transcription rate to increase What controls CAP? Cyclic AMP (cAMP) cAMP binds to CAP allosteric regulation Allows CAP to bind to the CAP binding site So..... what’s controlling cAMP cAMP production cAMP is produced from ATP by the enzyme adenylyl cyclase What controls adenylyl cyclase? Glucose! Lac operon High levels of glucose= low levels of cAMP Low levels of cAMP= low binding of CAP to the binding site Low binding of CAP to the binding site= reduced transcription of the lac genes High glucose slows transcription of the lac operon Why would this be adaptive Control of the lac operon Combination of positive and negative control allows for E.coli to quickly change their metabolism to exploit the best food sources