Download FROM DNA TO PROTEINS: gene expression Chapter 14 LECTURE

FROM DNA TO PROTEINS: gene expression
Chapter 14
LECTURE OBJECTIVES
What Is the Evidence that Genes Code for Proteins?
How Does Information Flow from Genes to Proteins?
How Is the Information Content in DNA Transcribed to Produce RNA?
How Is Eukaryotic DNA Transcribed and RNA Processed?
How Is RNA Translated into Proteins?
What Happens to Polypeptides after Translation?
ONE GENE ONE ENZYME IDEA
Phenotypic differences due to protein difference
Garrod studied ALKAPTONURIA
HA accumulated in blood due to an enzyme deficiency
MODEL ORGANISMS TO STUDY
Drosophila;
E. coli;
Neurospora crassa (common bread mold)
BEADLE & TATUM EXPERIMENT
one-gene, one-polypeptide relationship.
The gene-enzyme relationship has since been revised to the
Example: In hemoglobin, each polypeptide chain is specified by a separate gene.
Other genes code for RNA are not translated to polypeptides; some genes are
involved in controlling other genes.
THE CENTRAL DOGMA
The flow of information in cells
DNA→ RNA→ PROTEINS
REPLICATION
Two steps in making proteins:
Transcription
Translation
Remember RNA structure?
THREE KINDS OF RNA
Messenger RNA (mRNA)—carries copy of a DNA sequence to site of protein
synthesis at the ribosome
Transfer RNA (tRNA)—carries amino acids for polypeptide assembly
Ribosomal RNA (rRNA)—catalyzes peptide bonds and provides structure
FROM GENE TO PROTEIN
Exception to the central dogma:
retroviruses
transcription
We need:
DNA template
Nucleoside tri-phosphates (ATP, CTP, GTP, UTP)
RNA polymerase
RNA POLYMERASE
TRANSCRIPTION OCCURS IN 3 PHASES
INTITIATION
ELONGATION
TERMINATION
INITIATION
ELONGATION & TERMINATION
THE GENETIC CODE
Codon: A sequence of three bases—each codon specifies a particular amino acid.
Start codon: AUG—initiation signal for translation.
Stop codons: UAA, UAG, UGA—stop translation and polypeptide is released.
How was the code deciphered?
20 “code words” (amino acids) are written with only four “letters.”
Triplet code seemed likely: Could account for 4 × 4 × 4 = 64 codons.
DECIPHERING THE GENETIC CODE
THE GENETIC CODE
Is REDUNDANT
Not AMBIGUOUS
Is UNIVERSAL
Codons call for the same amino acids in all species
Exception: Chloroplast and Mitochondrion DNA
Genetic Code is the common language for evolution
Has remained the same
Facilitates genetic engineering
DIFFERENCES BETWEEN PROKARYOTES & EUKARYOTES
EUKARYOTIC GENES
Promoters and terminators
Non-coding genes: Introns
Coding sequences: Exons
Introns and exons appear in the primary mRNA transcript—pre-mRNA; introns
are removed from the final mRNA
EUKARYOTIC GENES
HOW INTRONS WERE DISCOVERED
in
Processing mRNA
In the nucleus, pre-mRNA is modified at both ends:
G cap is added at the 5′ end (modified guanosine triphosphate)—facilitates mRNA
binding to ribosome.
G cap protects mRNA from being digested by ribonucleases.
Poly A tail added at 3′ end.
AAUAAA sequence after last codon is a signal for an enzyme to cut the pre-mRNA;
then another enzyme adds 100 to 300 adenines—the “tail.”
Adding the g-cap
The snrnps (small nuclear ribonucleic particles) & SPLICEOSOME
In the disease β-thalassemia, a mutation may occur at an intron consensus
sequence in the β-globin gene—the pre-mRNA can not be spliced correctly.
Non-functional β-globin mRNA is produced.
Messenger rna leaving the nucleus
Through nuclear pores
Led by TAP proteins attaching to 5’ end
Unused mRNA stays in nucleus
TRANSLATION
Transfer RNA links information to the codons on m-RNA
Each t-RNA brings the specific amino acid
Transfer RNA must read codons correctly
Transfer RNA must bring the correct amino acid to m-RNA
Three functions of tRNA:
It binds to an amino acid, and is then “charged”
It associates with mRNA molecules
It interacts with ribosomes
Transfer rna
3′ end is the amino acid attachment site—binds covalently.
Anticodon: At the midpoint of the tRNA sequence—site of base pairing with mRNA.
Unique for each species of tRNA.
Wobble: Specificity for the base at the 3′ end of the codon is not always observed.
Example: Codons for alanine—GCA, GCC, and GCU—are recognized by the same
tRNA.
Wobble allows cells to produce fewer tRNA species, but does not allow the genetic
code to be ambiguous
CHARGING THE TRANSFER RNA MOLECULE
RIBOSOMES
Ribosomes have two subunits, large and small.
In eukaryotes, the large subunit has three molecules of ribosomal RNA (rRNA) and
49 different proteins in a precise pattern.
The small subunit has one rRNA and 33 proteins
RIBOSOMES
PARTS OF A RIBOSOME
A (amino acid) site binds with anticodon of charged tRNA
P (polypeptide) site is where tRNA adds its amino acid to the growing chain
E (exit) site is where tRNA sits before being released from the ribosome
Like transcription, translation also occurs in three steps:
Initiation
Elongation
Termination
INITIATION
An initiation complex forms—a charged tRNA and small ribosomal subunit, both
bound to mRNA.
In prokaryotes rRNA binds to mRNA recognition site “upstream” from start codon.
In eukaryotes the small subunit binds to the 5′ cap on the mRNA and moves until it
reaches the start codon.
INITIATION (start up codon on mRNA is AUG)
Elongation
The second charged tRNA enters the A site.
Large subunit catalyzes two reactions:
It breaks bond between tRNA in P site and its amino acid
Peptide bond forms between that amino acid and the amino acid on tRNA in the A
site
ELONGATION
Termination
Translation ends when a stop codon enters the A site.
Stop codon binds a protein release factor—allows hydrolysis of bond between
polypeptide chain and tRNA on the P site.
Polypeptide chain separates from the ribosome—C terminus is the last amino acid
added.
TERNMINATION
POLYRIBOSOME OR POLYSOME
POSTRANSLATIONAL ASPECTS
Protein modifications
Proteolysis: Cutting of a long polypeptide chain into final products, by proteases
Glycosylation: Addition of sugars to form glycoproteins
Phosphorylation: Addition of phosphate groups catalyzed by protein kinases—
charged phosphate groups change the conformation
POSTRANSLATIONAL MODIFICATIONS TO PROTEINS