Microbiology Procedure

Index >> Regulation of Protein Synthesis - Operon >> Promoter Genes

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Regulation of Protein Synthesis - Operon Structural Genes Operator Genes Promoter Gene Regulator Gene Control Mechanism - Induction Control Mechanism - Repression Inducible System Repressible System Negative Control of Protein Synthesis Positive Control of Protein Synthesis Mutation in the Operon

Promoter Genes

	Promoter Genes - The promoter gene is continuous with the operator gene. The complete nucleotide sequence of the control region of the lac operon of E. coli has been worked out by Gilbert and Maxam (1973), Maziels (1973) and Dickson et al (1975). This sequence extends from the termination codon (ACT on DNA= UGA on mRNA) of the structural gene, and includes the promoter and operator genes. The promoter region contains a sequence of two fold symmetry in which sections of DNA on either site of a particular point (axis of two fold symmetry) are symmetrical.
The sequences in these sections are 'palindromic'. The nucleotides of one strand go in one direction, while an identical sequence of nucleotides goes in the opposite direction in the other strand It has been suggested the these symmetrical regions of DNA may be recognized by proteins having symmetrically arranged subunits. The CRP site contains this section of two fold symmetry. The CRP site binds a protein called CRP (cyclic AMP receptor protein). This protein is essential for the binding of the enzyme RNA polymerase to the promoter. The CRP is a dimer with two subunits and a molecular weight of 45,000.
In E.coli the CRP combines with cyclic adenesine monophosphate (cAMP) to form a CRP-cAMP complex. This complex binds to the promoter. The CRP has a strong affinity for DNA, and this affinity increases considerably when the protein combines with cAMP. Binding of the CRP-cAMP complex to the promoter enhances RNA polymerase attachment to the promoter and thus increases transcription and protein synthesis. This is known as positive control
It has been found that in E.coli and certain phages the RNA polymerase binds to DNA at the promoter site and forms a stable complex. If such DNA molecules are treated with pancreatic DNase (an enzyme which breaks down DNA), the region of DNA forming the complex with RNA is protected from the action of DNase. This protected region is found to consist of 40-50 base pairs, and is not cleaved into smaller pieces by DNase.
	The RNA polymerase site has a small region of high A.T pairs, with high G.C' regions on either site. According to a model proposed by Pribnow (1975), the promoter region has three essential elements which are constant in position to each other. These elements are (i) a recognition sequence (ii) a binding sequence and (iii) an mRNA initiation site. The recognition sequence is outside the polymerase binding site, i.e. the region of DNA protected against the action of DNase. According to file model RNA polymerase first binds to DNA by forming a complex, with the recognition sequence. It then binds with the binding sequence to produce the preinitiation complex. The binding site consists of a sequence of seven bases. These are present in a constant location in the protected fragments. The sequence of this set of seven bases has been found to be almost constant in the bacteria and phages studied. It never differs by more than two bases from the following sequence: 5' T A TPi1A TG. RNA initiation site. The start of mRNA transcription takes place on one of two bases near the binding sequence. This starting site is included in the DNA fragment protected from DNase digestion. In the lac operon there is overlapping of promoter and operator sites. At least a part of the operator sequences appear in the polymerase binding site sequences In the promoter region, 20 base pairs to the left of the starting site of transcription, is a sequence S'CCGG. This is the substrate for an endonuclease enzyme isolated from Hemophilus parainfluenzae, and is know as the Hpa II- site. This site has been used as a reference point in studies of repressor and polymerase binding.

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Regulation of Gene Activity in Eukaryotes Frenster's Model of Gene Regulation Nonhistone Protein as Derepressors Britten Davidson Operon Operator Model Sensor Genes Producer Genes Integrator Genes Receptor Genes

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