Open promoter complexes

Various types of RNA polymerase-DNA binding complexes. A nonspecific complex is formed at any point along the DNA. The closed promoter complex is formed at a polymerase-binding site. Following the formation of the closed promoter complex, an open promoter complex is formed at the same site. 

The cr subunit is involved directly in promoter recognition. The complex lacking the polymerase core enzyme. To start transcription, the cr subunit directs the holoenzyme to a promoter site to form a binary complex in which there is a limited unwinding of the DNA duplex to generate an open promoter complex. This is the first step in the overall transcription cycle and is called template binding. 

The template-binding step involves interaction of the holoenzyme, through its cr subunit, with the promoter to give the open promoter complex, as already described. Initiation of the RNA chain can then proceed through the formation of the first phosphodiester bond between ATP (or GTP) and the next nucleotide defined by the template to yield a dinucleoside tetraphosphate ... 

First, the a subunit is needed for promoter binding and the formation of an open promoter complex. Subsequent to this, the /3 subunit (which will bind the inhibitor rifampicin) is essential for the formation of the first phosphodiester bond. 

The transition from the closed promoter complex (in which DNA is double helical) to the open promoter complex (in which a DNA segment is unwound) is an essential event in transcription. The stage is now set for the formation of the first phosphodiester bond of the new RNA chain. 

The answer is h. (Murray, pp 435-451. Scriver, pp 3-45. Sack, pp 1-40. Wilson, pp 101—120.) Promoter sites are initiation sites for transcription. Transcription starts when RNA polymerase binds to the promoter. It then unwinds the closed promoter complex, where DNA is in the form of a double helix, to form the open promoter complex in which about 17 base pairs of template DNA are unwound. RNA synthesis then begins with either a pppA or a pppG inserted at the beginning 5 -terminus of the new RNA chain, which is synthesized in the 5 to 3 direction. 

Initiation - After RNA polymerase has bound to a promoter and formed an open-promoter complex, the enzyme is ready to initiate synthesis of an RNA chain. One nucleoside triphosphate binding site on RNA polymerase is used during elongation. It binds any of the four common ribonucleoside triphosphates (rNTPs). Another binding site is used for initiation. It binds ATP and GTP preferentially. Thus, most mRNAs have a purine at the 5 end. 

RNA polymerase contacts - The -35 region and the -10 region, plus a few nucleotides upstream of -10, are the major contact points for RNA polymerase in an open-promoter complex. The RNA polymerase Dt subunit also makes contact in the -40 to -60 region. Figure 26.14 summarizes the results of these contacts for a promoter. Because there are two turns of the helix between the -35 and -10 regions, RNA polymerase is postulated to bind to DNA primarily on one side of the duplex. The data on nucleotide reactivity support this conclusion. 

Unwinding of DNA at promoter and formation of open promoter complex... 

Describe how topoisomerase I was used to determine the number of promoter DNA base pairs that are unwound upon the binding of RNA polymerase. Relate negative supercoiling to promoter efficiency. Distinguish between closed promoter and open promoter complexes. 

Bacterial RNA polymerase is a multisubunit enzyme of 480,000. The four major subunits—15, 5, a, and O (M = 150,000 160,000 36,500 and 86,000, respectively)—are present in the ratio 1 1 2 1, and the total complex is more correctly called the RNA polymerase holoenzyme. The o subunit is involved directly in promoter recognition. The complex without the o subunit is called the RNA polymerase core enzyme. To start transcription, a-helices in the O subunit simultaneously contact base pairs in the -10 and -35 regions of the promoter. This directs the holoenzyme to the promoter site to form a binary complex in which there is a limited unwinding of the DNA duplex to generate an open promoter complex. This is the first step in the overall transcription cycle and is called template binding. The sequence of events in the transcription cycle is illustrated in Fig. 9-3. 

Open-promoter complex A complex between RNA polymerase holoenzyme and a promoter that has undergone initial unwinding (has opened ) preparatory to the start of transcription. It is preceded by a much less stable closed-promoter complex, in which the promoter has not unwound, that may either fall apart or proceed to an open-promoter complex. 

Section 7.5). This enzyme-bound melted region is termed the open-promoter complex in which the enzyme is strongly bound to one chain of the DNA duplex called the sense or coding strand. This strand is transcribed into RNA. The other strand is not transcribed and is called the antisense strand. 

Upon the formation of an open-promoter complex, polyribonucleotide chain formation may commence. RNA polymerase contains two nucleotide-binding sites called the initiation and the elon-... 

DNA complex, while the A subunit binds to purified DNA. As a consequence of these divergent specificities, it has been proposed that the B subunit locates the specific site at which to bind to chromatin, while A may cause a conformational change in a localized region of chroma-tin-DNA, thereby providing sites for RNA polymerase to initiate mRNA synthesis. An alternate view, proposed by Yamamoto and Alberts (1976), is that binding of the receptor causes a cascade of histone acetylation, which makes the genome accessible to RNA polymerase. Perhaps the receptor, in a manner analogous to the cAMP receptor protein of prokaryotes, influences formation of an open-promoter complex with RNA polymerase (Krakow and Kumar, 1977). 

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