What does the word Promoter mean?
Download
1 / 86

What does the word Promoter mean? - PowerPoint PPT Presentation


  • 110 Views
  • Uploaded on

What does the word Promoter mean?. It is the place at which RNA Pol II binds. But the word is incorrectly used to describe Enhancers plus Promoter. Initiation by RNA Polymerase II. TFIID recognition site is TATAA. How often is this site found in the genome? 1/4 5.

loader
I am the owner, or an agent authorized to act on behalf of the owner, of the copyrighted work described.
capcha
Download Presentation

PowerPoint Slideshow about ' What does the word Promoter mean?' - geoff


An Image/Link below is provided (as is) to download presentation

Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author.While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server.


- - - - - - - - - - - - - - - - - - - - - - - - - - E N D - - - - - - - - - - - - - - - - - - - - - - - - - -
Presentation Transcript

What does the word Promoter mean?

It is the place at which RNA Pol II binds.

But the word is incorrectly used to describe

Enhancers plus Promoter.



Tfiid recognition site is tataa
TFIID recognition site is TATAA

How often is this site found in the genome? 1/45

Once every 1000 nucleotides 109 nucleotides or 106 times


Transient transfection

More Cells

But on a per cell

Basis expression levels of -gal

is about the same






Interrupted genes in eukaryotes exons and introns

Interrupted Genes in Eukaryotes: Exons and Introns

Most eukaryotic genes contain noncoding sequences called introns that interrupt the coding sequences, or exons. The introns are excised from the RNA transcripts prior to their transport to the cytoplasm.


Removal of intron sequences by rna splicing

Removal of Intron Sequences by RNA Splicing

The noncoding introns are excised from gene transcripts by several different mechanisms.



Splicing
Splicing

  • Removal of introns must be very precise.

  • Conserved sequences for removal of the introns of nuclear mRNA genes are minimal.

    • Dinucleotide sequences at the 5’ and 3’ ends of introns.

    • An A residue about 30 nucleotides upstream from the 3’ splice site is needed for lariat formation.


Types of intron excision
Types of Intron Excision

  • The introns of tRNA precursors are excised by precise endonucleolytic cleavage and ligation reactions catalyzed by special splicing endonuclease and ligase activities.

  • The introns of nuclear pre-mRNA (hnRNA) transcripts are spliced out in two-step reactions carried out by spliceosomes.


The spliceosome
The Spliceosome

  • Five snRNAs: U1, U2, U4, U5, and U6

  • Some snRNAs associate with proteins to form snRNAs (small nuclear ribonucleoproteins)



Logo for

a) Splice acceptor

b) Splice Donor

c) Initiator Met


AG/GT

CAG/NT

exon 1 intron 1 exon 2


Chapter 12 translation and the genetic code
Chapter 12Translation and the Genetic Code


Protein structure

Protein Structure

Proteins are complex macromolecules

composed of 20 (?)

different amino acids.


Amino acids
Amino Acids

  • Proteins are made of polypeptides.

  • A polypeptide is a long chain of amino acids.

  • Amino acids have a free amino group, a free carboxyl group, and a side group (R).


Peptide bonds
Peptide Bonds

  • Amino acids are joined by peptide bonds.

  • The carboxyl group of one amino acid is covalently attached to the amino group of the next amino acid.


Protein synthesis translation

Protein Synthesis: Translation

The genetic information in mRNA molecules is translated into the amino acid sequences of polypeptides according to the specifications of the genetic code.


The macromolecules of translation
The Macromolecules of Translation

  • Polypeptides and rRNA molecules Euk: 28S, 18S, 5.8S, 5S

  • Amino-acid Activating Enzymes

  • tRNA Molecules

  • Soluble proteins involved in polypeptide chain initiation, elongation, and termination



Ribosomes
Ribosomes

What does “S” mean?

Why do sizes get bigger?


The nucleolus
The Nucleolus

  • In eukaryotes, the nucleolus is the site of rRNA synthesis and ribosome assembly




Rrna genes
rRNA Genes Mammals

  • rRNA Genes in E. coli

    • Seven rRNA genes distributed among three sites on the chromosome

  • rRNA Genes in Eukaryotes

    • rRNA genes are present in hundreds to thousands of copies

    • The 5.8S-18S-28S rRNA genes are present in tandem arrays in the nucleolar organizer regions of the chromosomes.

    • The 5S rRNA genes are distributed over several chromosomes.


Transfer rnas trnas
Transfer RNAs (tRNAs) Mammals

  • tRNAs are adapters between amino acids and the codons in mRNA molecules.

  • The anticodon of the tRNA base pairs with the codon of mRNA.

  • The amino acid is covalently attached to the 3’ end of the tRNA.

  • tRNAs often contain modified nucleosides.



Inosine Mammals



Specificity of trnas
Specificity of tRNAs Mammals

  • tRNA molecules must have the correct anticodon sequence.

  • tRNA molecules must be recognized by the correct aminoacyl-tRNA synthetase.

  • tRNA molecules must bind to the appropriate sites on the ribosomes.




Stages of translation
Stages of Translation enzyme)

  • Polypeptide Chain Initiation

  • Chain Elongation

  • Chain Termination


Translation initiation in e coli
Translation Initiation in enzyme)E. coli

  • 30S subunit of the ribosome

  • Initiator tRNA (tRNAMet)

  • mRNA

  • Initiation Factors IF-1, IF-2, and IF-3

  • One molecule of GTP

  • 50S subunit of the ribosome



Translation initiation in eukaryotes
Translation Initiation in Eukaryotes enzyme)

  • The amino group of the methionine on the initiator tRNA is not formylated.

  • The initiation complex forms at the 5’ terminus of the mRNA, not at the Shine-Dalgarno/AUG translation start site.

  • The initiation complex scans the mRNA for an AUG initiation codon. Translation usually begins at the first AUG.

  • Kozak’s Rules describe the optimal sequence for efficient translation initiation in eukaryotes.


Polypeptide chain elongation
Polypeptide Chain Elongation enzyme)

  • An aminoacyl-tRNA binds to the A site of the ribosome.

  • The growing polypeptide chain is transferred from the tRNA in the P site to the tRNA in the A site by the formation of a new peptide bond.

  • The ribosome translocates along the mRNA to position the next codon in the A site. At the same time,

    • The nascent polypeptide-tRNA is translocated from the A site to the P site.

    • The uncharged tRNA is translocated from the P site to the E site.



Polypeptide chain termination
Polypeptide Chain Termination Ribosomes

  • Polypeptide chain termination occurs when a chain-termination codon (stop codon) enters the A site of the ribosome.

  • The stop codons are UAA, UAG, and UGA.

  • When a stop codon is encountered, a release factor binds to the A site.

  • A water molecule is added to the carboxyl terminus of the nascent polypeptide, causing termination.




The genetic code

The Genetic Code Ribosomes

The genetic code is a nonoverlapping code, with each amino acid plus polypeptide initiation and termination specified by RNA codons composed of three nucleotides.


Properties of the genetic code
Properties of the Genetic Code Ribosomes

  • The genetic code is composed of nucleotide triplets.

  • The genetic code is nonoverlapping. (?)

  • The genetic code is comma-free. (?)

  • The genetic code is degenerate. (yes)

  • The genetic code is ordered. (5’ to 3’)

  • The genetic code contains start and stop codons. (yes)

  • The genetic code is nearly universal. YES :)


A triplet code
A Triplet Code* Ribosomes





Evidence of a triplet code in vitro translation studies
Evidence of a Triplet Code: frame.*In Vitro Translation Studies

  • Trinucleotides were sufficient to stimulate specific binding of aminoacyl-tRNAs to ribosomes.

  • Chemically synthesized mRNAs containing repeated dinucleotide sequences directed the synthesis of copolymers with alternating amino acid sequences.

  • mRNAs with repeating trinucleotide sequences directed the synthesis of a mixture of three homopolymers.




The genetic code1
The Genetic Code frame.*

  • Initiation and termination Codons

    • Initiation codon: AUG

    • Termination codons: UAA, UAG, UGA

  • Degeneracy: partial and complete

  • Ordered

  • Nearly Universal (exceptions: mitochondria and some protozoa)


Key points
Key Points frame.*

  • Each of the 20 amino acids in proteins is specified by one or more nucleotide triplets in mRNA. (20 amino acids refers to what is attached to the tRNAs!)

  • Of the 64 possible triplets, given the four bases in mRNA, 61 specify amino acids and 3 signal chain termination. (have no tRNAs!)


Key points1
Key Points frame.*

  • The code is nonoverlapping, with each nucleotide part of a single codon, degenerate, with most amino acids specified by two to four codons, and ordered, with similar amino acids specified by related codons.

  • The genetic code is nearly universal; with minor exceptions, the 64 triplets have the same meaning in all organisms. (this is funny)


Do all cells/animals make the same frame.*

Repertoire of tRNAs?


The wobble hypothesis base pairing involving the third base of the codon is less stringent
The Wobble Hypothesis: frame.*Base-Pairing Involving the Third Base of the Codon is Less Stringent.



Suppressor mutations
Suppressor Mutations frame.*

  • Some mutations in tRNA genes alter the anticodons and therefore the codons recognized by the mutant tRNAs.

  • These mutations were initially detected as suppressor mutations that suppressed the effects of other mutations.

  • Example: tRNA mutations that suppress amber mutations (UAG chain-termination mutations) in the coding sequence of genes.



Translation of an amber uag mutation in the absence of a suppressor trna
Translation of an frame.*amber (UAG) Mutation in the Absence of a Suppressor tRNA


Translation of an amber mutation in the presence of a suppressor trna
Translation of an frame.*amber Mutation in the Presence of a Suppressor tRNA

Note it is amber su3…why?????????


Translation of an amber mutation in the presence of a suppressor trna1
Translation of an frame.*amber Mutation in the Presence of a Suppressor tRNA

If there was a single tRNATyr gene, then could one

have a amber supressor of it?


Historical comparisons
Historical Comparisons frame.*

  • Comparison of the amino acid sequence of bacteriophage MS2 coat protein and the nucleotide sequence of the gene encoding the protein (Walter Fiers, 1972).

    Was this first????

  • Sickle-cell anemia: comparison of the sequence of the normal and sickle-cell alleles at the amino acid level and at the nucleotide level.


Are the proteins produced a pure reflection of the mrna sequence
Are the proteins produced a pure reflection of the mRNA sequence????

tRNA environment, protein modifications post-translationally


To Know for Exam sequence????

RNApol II

TATAA

CCATGG (Nco I site and Kozak Rule)

ATG

AGGT….splice

GT……………A………polypyrimidine AG

PolyA recog sequence

AATAAA

The Reasons why ATG is a single codon

and TGG is a single codon.


ad