BIOL10221 Molecular Biology Lecture Notes PDF

BIOL10221 “Molecular Biology” Dr David Boam Module 4 - From DNA to RNA Lecture 3: Gene expression in Prokaryotes: The Lac and Trp operon  Control of Gene Expression How do polymerases know when to transcribe a gene? Lac operon Trp operon...

BIOL10221 “Molecular Biology” Dr David Boam Module 4 - From DNA to RNA Lecture 3: Gene expression in Prokaryotes: The Lac and Trp operon  Control of Gene Expression How do polymerases know when to transcribe a gene? Lac operon Trp operon Operon - cluster of genes transcribed by the same promoter that gives rise to a polycistronic mRNA - genes usually related Lecture 4/3 lac and trp operon The Lac Operon hydrolyses lactose to galactose and glucose Lecture 4/3 lac and trp operon  The lac operon No lactose - lac operon is repressed. I CRP P O Z Y A Repressor binds to O preventing RNA polymerase from clearing promoter (P). Allolactose binds to repressor - dissociates from the Operator lactose is an inducer - (in fact de- repression) It is under negative feedback control. Lecture 4/3 lac and trp operon  Cis-acting elements and trans-acting factors Trans-acting gene Trans-acting factor Cis-acting - will only regulate DNA to which it is directly joined “dominant” Promoter (cis-acting sequence) Trans-acting will regulate genes anywhere mostly protein transcription factors recessive (mostly) Lecture 4/3 lac and trp operon Cis-acting and trans-acting mutations - dissecting the lac operon lacOc lacI- X X Mutated operator Mutated repressor  constitutive  constitutive I+ I+ I- X X Introduction of I+ Introduction of I+  no effect  complements cis-dominant  trans- Lecture 4/3 lac and trp recessive operon  Lac operon - Summary Repressor binds operator – inhibits initiation Repressor activity regulated by lactose Mutations in regulatory elements – Cis acting – Cannot be complemented Mutations which alter activity of regulatory proteins – Trans-acting – May affect many genes – Can be complemented Lecture 4/3 lac and trp operon  Catabolite repression Diauxic growth Glucose exhausted, growth on lactose Glucose+ growth lactose b gal lag Activation of operon Several operons allowing use of alternate carbon sources are repressed by glucose and only become active when all glucose is used up. Preferential use of glucose This occurs by catabolite repression. Lecture 4/3 lac and trp operon  Catabolite repression - the role of CRP I O Z Y A P CRP CRP=Catabolite Repressor Protein cAMP (sometimes called CAP = Catabolite Activator Protein) RNA polymerase requires CRP cAMP binds to CRP permitting DNA binding High glucose = low cAMP….so lac operon is off, even if lactose is available Low glucose = high cAMP ….. so lac operon is on if lactose present Glucose preferentially used as a carbon source Lecture 4/3 lac and trp operon  The Trp Operon ontains genes for synthesis of amino acid tryptophan Trp repressor - binds to operator in presence of tryptophan and prevents transcription Lecture 4/3 lac and trp operon  Keywords/concepts Trp operon – REPRESSED by tryptophan (biosynthetic PRODUCT) Compare – to lac operon INDUCED by lactose (substrate) Products of biosynthetic operons repress their operons Substrates for metabolism induce their operons Lecture 4/3 lac and trp operon  Intended learning objectives By completion of this lecture you should be able to Define and describe the functional components of a typical operon Understand the mechanism of regulation of the lac operon by lactose and glucose Understand the concept of negative feedback regulation Understand the mechanism of catabolite repression Understand the following general concepts and how they apply to mechanism of regulation by operons – DNA-protein interactions – protein-protein interactions – protein-small molecule interactions – conformational change – inducers/co-repressors Lecture 4/3 lac and trp operon