Gene regulation Gene regulation Gene involved in a

Gene regulation

Gene regulation • Gene involved in a metabolic pathway are often present in a linear array called operon • Operon can be regulated by a single promoter or regulatory region • Cistron is the smallest unit of genetic expression • A single m. RNA that encodes more than one separately translated protein is called poly cistronic m. RNA

• Induction: - increased synthesis of proteins or enzymes in response to certain signals(inducers) • It is turning “on” the switch of the gene • Inducible gene: -one whose expression increases in response to an ‘inducer’ or ‘activator’ • Repression is the turning “off “ the switch of the gene

Ø There are two types of gene regulation: v. Negative regulation v. Positive regulation • Negative regulation: üDefault state is “on” and ‘transcription’ takes place until it is turned “off” by a “repressor” üIt may be either : Inducible Repressible

Inducible transcription ü A repressor DNA –binding protein keeps transcription in the “off” state ü In presence of inducers , the repressor binds with inducer and losses its DNA binding capability , and transcription occurs Repressible transcription ü Default state is “on" until an active repressor is formed to turn it “off”

ü The regulatory protein here is called as “aporepressor” and has no DNA -binding activity of its own ü The active repressor is formed by combination of the aporepressor and a small molecule known as the co-repressor ü Presence of co-repressor result in the cessation of transcription

• Positive regulation • Default state is “off” and binding with a regulatory protein is required to turn it “on” • Regulatory protein is called “transcriptional activator protein”

Auto regulation: - • Protein product of gene regulates its own transcription • Two types: -Negative auto regulation Positive auto regulation v Negative auto regulation: ü Protein inhibits transcription and high concentration of the protein result in less transcription of the m. RNA v Positive auto regulation: ü The protein stimulate transcription, as more is made, transcription increases to the maximum rate

Lac operon system of gene regulation Ø Francois Jacob and Jacques Monad in 1961 described the operon model Ø Based on the observation of lactose metabolism in Escherichia coli Ø Cells are grown in glucose medium do not contain beta galactosidase (lactase) , but when transferred to a medium containing only lactose , then the enzyme level in the cell increases several thousand folds

Features of lac operon are: - Ø Operon include : structural gene , control elements , regulator/inhibitor genes , promoter and operator areas

Two genes are required for lactose utilization: - Ø Lac Z gene : encodes β – galactosidase Ø Lac Y gene: encodes lac permease Ø Another gene Lac A encodes thiogalactoside transacetylase Ø These three genes are called structural genes and present as continuous segments of DNA Ø Transcription of these gene start from a common promoter (P) located close to Z gene

Ø Synthesis of lac m. RNA is determined by activity of specific regulatory protein which is encoded by Lac I gene Ø Lac repressor molecule is a tetramer having high affinity for the operator locus Ø When attached to the operator locus , lac I repressor molecule prevents transcription of distal structural genes(Lac Z, Lac Y, Lac A) by interfering the binding of RNA polymerase

Ø An inducer is needed to initiate transcription of m. RNA. The inducer of Lac operon is allolactose Ø It binds to the repressor and alter the three dimensional structure that is unable to bind to the operator Ø Lac I repressor molecule has high affinity for the inducer Ø It induce a conformational change in the structure of repressor and cause it to dissociate from operator DNA

Ø DNA dependent RNA polymerase now bind to the coding strand at the promoter site and transcription will begin Ø Thus an inducer derepresses the lac operon and allow transcription of structural gene

Ø Inorder for the RNA polymerase to bind at the promoter site most efficiently, there must be presence of “catabolic gene activator protein”(CAP) to which c. AMP is bound Ø The bacterium accumulate c. AMP only when it is starved for a source of carbon Ø In presence of glucose or glycerol, c. AMP saturated CAP is lacking, so DNA dependent RNA polymerase cannot initiate transcription of the lac operon at the maximum rate

Ø In presence of CAPc. AMP complex, DNA dependent RNA polymerase binds to DNA just upstream of the promoter site, transcription occurs at maximum level

Ø A lactose analog that is capable of inducing the lac operon while not itself serving as a substrate for galactosidase is called gratuitous inducer. Ø E. g: isopropyl thiogalactoside(IPTG) Ø Addition of lactose or IPTG to bacteria growing on a poorly utilized carbon source(eg: succinate) result in prompt induction of lac operon enzymes β-

Tryptophan (trp) operon Ø Tryptophan operon is responsible for the production of amino acid tryptophan Ø The enzyme coding sequence in DNA has a ‘promoter’, ’operator’, and two regions called the ‘ leader (trp l) ’ and the ‘attenuator (trp a)’ Ø Another gene ‘trp R’ encoding a repressor is located some distance from this gene

Ø Trp operon contain structural genes for enzymes that synthesis tryptophan and it occurs in five steps each require a particular enzyme Ø The genes are: -trp E , trp D , trp C , trp B, trp A Ø trp E coding region is first translated Ø The product of trp R gene is the ‘aporepressor protein’ Ø Tryptophan act as a co–repressor which binds with the trp aporepressor to form the active repressor

Ø Binding of repressor to trp operator shuts off synthesis Ø When there is not enough tryptophan, the Apo repressor adopt three dimensional conformation unable to bind with trp operator and the operon is transcribed Ø When tryptophan is present at high concentration, it binds to aporepressor and forms and cause it to change conformation into active repressor. It binds to the operator and prevent transcription

Attenuation Ø When tryptophan is present in growth medium , but not enough to sustain optimal growth, under these condition it is advantageous for cell to synthesis tryptophan , but at less than the maximum possible rate Ø The amount of transcription in the depressed state is controlled quantitatively by the concentration of tryptophan in the cell Ø This regulatory mechanism is called attenuation

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