Elementary plant Biochemistry and Biotechnology (2+1)

Binding of RNA polymerase

• E. coli RNA polymerase consists of five subunits – two identical α subunits and one each types of β, β’ and σ. The σ subunit dissociates from the enzyme during the elongation stage of RNA polymerization. The term core enzyme is used to describe the σ.-free unit, α2ββ. The complete enzyme, α2ββ is called the holoenzyme.


• RNA polymerase is sufficiently large that it can come into contact with many DNA bases simultaneously. The first step in transcription is the binding of RNA polymerase to a DNA molecule. Binding occurs in particular regions called promoters, which are sequences in which several interactions occur.


• Several events occur at a promoter: RNA polymerase recognize a specific DNA sequence, attach in a proper conformation, locally open the DNA strands in order to gain access to the bases to be copied and then initiate synthesis. These events are guided by the base sequence of the DNA, the polymerase σ subunit (without which the promoter is not recognized).


• The specific binding region in DNA is in a region 5 –10 bases prior to the left of the first base copied into mRNA. This region is called as Pribnow box. All sequences found in Pribnow boxes are considered to be variants of a basic sequence TATAAT, hence they are called as TATA box. The first base transcribed was chosen as a reference point and numbered +1. The direction of transcription was called downstream; all upstream bases, which are not transcribed, were given negative numbers starting from the reference. Thus, the Pribnow box is enclosed between –21 and –4 depending on the particular promoter.


• There is a second important region, to the left of the Pribnow box, whose sequences in different promoters have common features. This six base sequence which is called as the “–35 sequence “has a consensus TTGACA, may be initial site of binding of the enzyme, when the sequence is present. The Pribnow box is thought to orient RNA polymerase, such that synthesis proceeds from left to right and to be the region at which the double helix opens to form the open- promoter complex. This highly stable complex is the active intermediate in chain initiation.


• The DNA double helix in an open-promoter complex is locally unwound, starting about 10 bp from the left end of Pribnow box and extending about 20 bp past the position of the first transcribed base. It seens that RNA polymerase itself induces this unwinding and undergoes a conformational change itself in so doing. This melting is necessary for pairing of incoming ribonucleotides.