Promotor Recognition

The recognition sequences of regulatory proteins may overlap not only the promotor site, but can also be found in the immediate vicinity of the a promoter. The sequence elements are relatively simple and often include only one binding site for regulatory proteins. 

At low Trp concentration, the Trp repressor is mainly in the unbound, inactive form. The free form of the Trp repressor binds with a ca. 104-fold lower affinity to the recognition sequence than that of the Trp-bound form. The promotor remains free under these conditions, and transcription of the genes for Trp biosynthesis can occur. The shutting on and off of the Trp operon is based on the disparate DNA affinities of the free and Trp-bound repressor. 

The RNA polymerase of E. coli possesses with its subunit construction (afjfj a) a simple structure in comparison to eucaryotic RNA polymerases. The sigma factor is only required for the recognition of the promotor and the subsequent formation of a tight complex. After the incorporation of the first 8-10 nucleotides into the transcript, the sigma factor dissociates from the holoenzyme, and the remaining core enzyme carries out the rest of the elongation. 

Key elements of dependent promotors are the TATA box, with the consensus sequence TATAAT 10 bp upstream from the transcription initiation site (pos. -10), and the sequence TTGACA at the position -35 (Fig. 1.18). Both sequences are necessary for the recognition of the promotor by a70. Structural analysis of the Thermus aquaticus RNA polymerase holoenzyme bound to DNA shows that all sequence-specific contacts with the core promotor are mediated by the sigma subunit (Murakami et al. 2002). This archaeal RNA polymerase has a subunit structure (a ji/i oxr) similar to that of the eubacterial enzyme. The intervening sequences, as well as other upstream sequences, can also influence the efficiency of transcription initiation. It is not possible to define consensus sequences at these positions. An optimal dependent promotor can be defined as a sequence with the - 35 hexamer as well as the -10 hexamer 17 bp away. The latter lies 7 bp upstream from the transcription initiation site. 

Substrate recognition and selection of the JNK/SAPK and p38 proteins (and also the ERK proteins) are mediated both by specific docking sites and by the nature of the amino acids surrounding the phosphoacceptor site. For the transcription factor substrates, specific docking domains have been identified that are loacted at a distance from the phosphorylation sites in the transactivation domain. These docking sites serve to increase the selectivity and specificity of phosphorylation, and they are used for recruitment of MAPK kinases into protein complexes at promotors, where they can phosphorylate other regulatory transcriptional proteins. 

The problem of recognition apparently is important both in replication and transcription, although it is rarely discussed in physical terms. The question why polymerases start and stop their action at a given point (to one nucleotide exactly ) is completely open. Recent progress in the elucidation of the sequences of RNA polymerase promotor sites shows some degree of homology between these promotors (41). There must be initiation and termination signals for all these mechanisms, as well as for their control (operon, repressor sites, etc.) (42). 

---