What is the flow of information through the cell?

Major grooveMinor groove

Double helix - antiparallel polymers

5’

3’

A

CG

T

Purine Pyrimidine

06_12_asymmetrical.jpg

Transcription:• dsDNA template • Nucleotides (ACGU)

make ssRNA• Need to separate

strands.• Nucleotides added to

free 3’ OH (5’3’)

Classes of RNA…

mRNA rRNA tRNA

…also snRNA and microRNA

DNA

mRNA

polypeptide

coding sequence

untranslated regions5’ UTR 3’ UTR

Prokaryotes

promoter(not transcribed)

RNA Polymerase

Ribosome

DNA

pre mRNA

polypeptide

promoter(not transcribed)

coding sequence

3’ UTR

Eukaryotes

mRNA

untranslated regions5’ URT

RNA Polymerase

Ribosome

Bacterial Promoter Elements

• Transcription start = +1• Consensus sequence = –35; TTGACA, recognized by • Pribnow box = -10, TATAAT; determines +1• Terminator sequence: where polymerase stops

Initiation of transcription in prokaryotes

Initiation of eukaryotic transcription by RNA Pol II (mRNA)

TF = transcription factor (compare with prokaryotic sigma factor)

Eukaryotic mRNA:

• 5’ cap,• 5’ UTR• coding region• 3’ UTR• 3’ poly-A tail

mRNA processing in Eukaryotes

5’ cap added

Remove 3’ end

Poly-A tail added

Introns removed

Exons joined

DNA RNA Protein

transcription translationreplication

Gene cloning - making lots of copies...1. Make a “library” of small pieces of DNA (2 types)2. Find the one piece you want3. Insert it into a “vector”4. Grow it in a new organism (bacteria, euk. cells)

Isolate DNA, fragment with RE

Isolate mRNA, convert to cDNA with reverse transcriptase

Genomic library

cDNA library

07_37_Protein.produc.jpgOverview of gene expression in eukaryotes

07_28_ribosome.jpg

07_26_2_adaptors.jpgTwo adapters link an amino acid to a codon

07_32_initiation.jpg

Intiation of translation in Eukaryotes

07_33_mRNA.encode.jpg

Intiation of translation in Prokaryotes

07_30_3_step_cycle.jpgElongation of proteins

4

5

07_34_stop codon.jpg

Termination of translation

Mutations…• Frameshift: Adding or removing 1 or 2 nucleotides results

in changes the reading frame from that point on.

• Nonsense: Changing an amino acid codon to a stop codon results in truncated proteins

• Missense: Changing an amino acid codon to one encoding a different amino acid - effect depends on type of amino acid and where in the protein.

04_03_20 amino acids.jpg

Side chains interact via all of the noncovalent bonds

Primary structure (1°) of a protein:

Arabidopsis -glucosidase (single letter codes)

MSSLHWFPNIFIVVVVFFSLRSSQVVLEEEESTVVGYGYVVRSVGVDSNRQVLTAKLDLIKPSSVYAPDIKSLNLHVSLETSERLRIRITDSSQQRWEIPETVIPRAGNHSPRRFSTEEDGGNSPENNFLADPSSDLVFTLHNTTPFGFSVSRRSSGDILFDTSPDSSDSNTYFIFKDQFLQLSSALPENRSNLYGIGEHTKRSFRLIPGETMTLWNADTGSENPDVNLYGSHPFYMDVRGSKGNEEAGTTHGVLLLNSNGMDVKYEGHRITYNVIGGVIDLYVFAGPSPEMVMNQYTELIGRPAPMPYWSFGFHQCRYGYKNVSDLEYVVDGYAKAGIPLEVMWTDIDYMDGYKDFTLDPVNFPEDKMQSFVDTLHKNGQKYVLILDPGIGVDSSYGTYNRGMEADVFIKRNGEPYLGEVWPGKVYFPDFLNPAAATFWSNEIKMFQEILPLDGLWIDMNELSNFITSPLSSGSSLDDPPYKINNSGDKRPINNKTVPATSIHFGNISEYDAHNLYGLLEAKATHQAVVDITGKRPFILSRSTFVSSGKYTAHWTGDNAAKWEDLAYSIPGILNFGLFGIPMVGADICGFSHDTTEELCRRWIQLGAFYPFARDHSSLGTARQELYLWDSVASSARKVLGLRMRLLPHLYTLMYEAHVSGNPIARPLFFSFPQDTKTYEIDSQFLIGKSIMVSPALKQGAVAVDAYFPAGNWFDLFNYSFAVGGDSGKHVRLDTPADHVNVHVREGSIVAMQGEALTTRDARKTPYQLLVVASRLENISGELFLDDGENLRMGAGGGNRDWTLVKFRCYVTGKSVVLRSEVVNPEYASKMKWSIGKVTFVGFENVENVKTYEVRTSERLRSPRISLIKTVSDNDDPRFLSVEVSKLSLLVGKKFEMRLRLT

Secondary structure (2°)

-helix

H-bonds between CO and NH of backbone.(No R-groups involved)

Secondary structure

-sheet

H-bonds between CO and NH of backbone.(No R-groups involved)

-helix

-sheet

loops and turns

disulfide bridge

Tertiary structure - the entire polypeptide

ribonuclease

Quaternary structure - multiple subunits e.g. hemoglobin

04_20_protein domains.jpgcytochrome b562

lactic (lactate)dehydrogenase immunoglobulin

light chain

Domains - discrete modules within tertiary structure that fold independently and have a specific function.

4 domains of phospholipase C

Motif - a recurring substructure barrel e.g. -amylase

Motif - a recurring substructurecoiled coil e.g. myosin

How do proteins get to their folded state?

unfolded nativeconformation

04_21_Serine proteases.jpg

Proteins with different functions may have similar shape - members of a family with a common ancestor.

04_22_protein subunit.jpg