Shine-Dalgarno Motif

• Ribosome binding site located about 13 bases upstream of AUG start codon

• SD sequence is: 5’-AGGAGGU-3’

• Middle GGAG is more highly conserved

• Degree of match is positively correlated with translation rate

Assignment (due 9/9/04)

• Write perl code to detect Shine-Dalgarno motifs in E. coli DNA sequences.

• I will give you about 15 files, each has a single gene and one SD motif. (Ecoli.1, Ecoli.2, etc. in “examples”.

• You do not know which ORF to use (except that it is in the forward direction, so only 3 choices).

• S-D will be 6 bp (±5) upstream of ATG.• Perfect matches are the exception, not the rule.• Which ATG is a start codon? (Hint: gene will be at least 250 bp

long, so if you encounter a stop codon early, it is not a gene. If you do not encounter a stop codon ever, it is not a gene.)

Eukaryotes (eucaryotes)

Cells with a nucleus

(us)

Gene expression: eukaryotes vs. prokayotes

RNA polymerase II promoters

• BRE - TFIIB recognition element

• TATA box• INR - Intiator region

Py.Py.A*.N.T/A.Py.Py• (Py = pyramidine C/T)• DPE - downstream promoter

element• Other elements 100-300 bp

upstream CAT box and GC box

Initiation of transcription

• TFIIA, B, C … general transcription factors

• TBP - TATA binding protein

• Over 100 proteins involved

• TFIIH pulls DNA apart

• Then transcription factors are released so that transcription can occur

• Similar to sigma factor in prokaryotic transcription

In vivo is even more complex; note long distance regulatory proteins

Recognition sequences for most common splice (Y =C/U(T), R=A/G)

“Lariat” mechanism for removing intron

GT(U) - AG rule

• Most common splice mechanism

• U1, U2, U4, U5, U6 are snRNPs (small nuclear ribonucleoproteins)

• snRNPs are made up of snRNAs and proteins

• Assembly of snRNPs and other proteins is called a spliceosome

Alternative splicing!