1. DNA-Protein Interactions & Complexes

6. Prokaryotic promoter Consensus sequence is not present in majority of prokaryotic promoters. Sequence motifs encompassing 6-10 nucleotides are degenerate; Probability of finding similar sequences in regions other than promoters is quite high. Harley and Reynolds (1987) Nucleic Acids Res. 15(5): 2343-2361. 6

7. Sequence logo for B. subtilis, E. coli and M. tuberculosis promoter sequences No sequence conservation 7

8. Does this indicate that there are other signals which help in positioning RNA polymerase? Hence analysis of structural properties of a DNA sequence to locate signals that are: Relevant to transcription from a functional/mechanistic/structural point of view. Unique to the promoter sequences and can be used to differentiate between promoter and non-promoters. Can be predicted from a given sequence. For example: 1) DNA STABILITY (Ability of DNA to Open up) 2) DNA CURVATURE (Intrinsically curved DNA structure) 3) DNA BENDABILITY (Ability of DNA to bend)

9. An important step in transcription is the formation of an open complex which involves strand separation of DNA duplex upstream of the transcription start site (TSS) This separation takes place without the help of any external energy. Hence evaluating stabilities of promoter sequences may give some clues. Why Stability?

10. Stability of base paired dinucleotides SantaLucia J (1998) Proc. Natl. Acad. Sci. USA 95(4):1460-1465. based on Tm (melting temp data) on a collection of 108 oligonucleotide duplexes.

11. A representative free energy profile for 1000nt long E. coli promoter sequence

14. DNA bendability Protein DNA

16. TATA- containing promoters are less stable (fig a), highly bendable and more curved (fig d) as compared to TATA-less promoters. The average free energy, Dnase1 sensitivity, Nucleosome positioning preference and curvature profiles of S.cerevesiae promoters w.r.t TSS. TATA- containing promoters are less stable (fig a), highly bendable (fig b) and more curved (fig d) as compared to TATA-less promoters. The nucleosome positioning preferences (fig c) are only slightly different. TATA-containing promoters show different structural properties from TATA-less promoters

17. Different DNA binding helix turn helix protein motifs