From DNA to Protein
DNA is just instructions
Need to use the instructions
2 processes
Transcription—molecules of RNA formed from DNA
Translation—RNA shipped from nucleus to cytoplasm to be used in polypeptide
DNA to RNA
3 classes of DNA
Messenger RNA: carries blueprint to ribosome
Ribosomal RNA: combines with proteins to form ribosomes upon polypeptides are assembled
Transfer RNA: brings correct amino acids to ribosome and pairs up with mRNA code for that amino acid
RNA
Different sugar: ribose
1 different base: uracil instead of thymine
Cellular distribution: nucleus and cytoplasm
Transcription compared to Replication
Only 1 region of DNA strand is used as a template
RNA polymerase instead of DNA polymerase
Result of transcription is single-stranded RNA
RNA
Transcription begins when RNA polymerase binds to promoter region
Moves along gene until the end of the gene
Copies code
Sustitutes uracil (U) for thymine base
RNA
After copying info, RNA is modified
Cap is attached to 5’end: start
Tail is attached at end (poly A): finish
RNA is edited
Introns—no code—removed
Exons—code—used to sequence amino acids
The Structure of Proteins
Proteins are made from subunits called amino acids
Hundreds of thousands of different proteins made by all living things are remarkably similar in their construction
All proteins in living things are assembled from only 20 different amino acids
The Structure of Proteins
These 20 amino acids are strung together in different orders and to different lengths to make different proteins
The English alphabet contains a similar number of letters, but we can form an infinite number of sentences with these letters
The Structure of Proteins
A linear chain of amino acids forms a polypeptide
A polypeptide chain can be more than 20 amino acids in length
Only 20 different amino acids are used
Some amino acids are used multiple times
The Structure of Proteins
One or more polypeptide chains are folded into a single protein
Each protein has a particular three-dimensional shape
“Protein conformation”
This shape is stabilized by chemical bonds
Covalent, ionic, and hydrogen bonds
The shape of a protein is critical to its function
4.3 A Closer Look at Transcription
DNA
Polymer of deoxynucleotides
Sugar
Deoxyribose
Bases
G, A, C, and T
Phosphates
RNA
Polymer of nucleotides
Sugar
Ribose
Bases
G, A, C, and U
Phosphates
A Closer Look at Transcription
The enzyme RNA polymerase unwinds a region of the DNA double helix
(RNA polymerase is actually an enzyme complex, consisting of a group of enzymes)
The two strands of the double helix are separated
Single-stranded DNA is exposed
A Closer Look at Transcription
RNA polymerase assembles complementary RNA nucleotides
DNA:RNA base pairing similar to DNA:DNA
Recall base pairing from Chapter 13
G:C
C:G
A:U
T:A
A Closer Look at Transcription
The completed portion of the RNA molecule separates from the DNA
DNA double helix is rejoined in this region
RNA polymerase moves along and unwinds more of the double helix
New (untranscribed) regions of single-stranded DNA are exposed
A Closer Look at Transcription
Upon completion of the RNA transcript
The mRNA transcript is released from the DNA
The DNA strands rejoin
The DNA and enzyme are unchanged
A new mRNA molecule has been produced
Transcription
Transcription video
A Closer Look at Transcription
Transcription takes place in the nucleus
Translation takes place in the cytoplasm
Following the production of an mRNA molecule, it must be transported to the cytoplasm
Transport is through a nuclear pore
14.4 A Closer Look at Translation
Translation requires many players
mRNA
Ribosomes
Transfer RNAs (tRNAs)
Amino acids
A Closer Look at Translation
mRNA molecules
Groups of three consecutive nucleotides are the functional units within mRNA molecules
“Codons”
Each codon corresponds to a specific amino acid
e.g., AUG methionine
e.g., UUU phenylalanine
Cracking the Genetic Code
The universal nature of the genetic code is useful in many ways
Knowing a gene’s DNA sequence tells us the protein’s amino acid sequence
Knowing a protein’s amino acid sequence tells us much about the gene’s DNA sequence
Genes from one organism can function in another organism: Makes “biotechnology”possible
A Closer Look at Translation
tRNA molecules
“Transfer RNA”
Encoded by genes
Functional as RNA molecules
Not translated into proteins
“Translates”information from mRNA to protein
A Closer Look at Translation
tRNA molecules
One region binds to the mRNA molecule
“Anticodon”
Base pairs with mRNA codon
Another region is linked to a specific amino acid
A Closer Look at Translation
Ribosomes
Organelles
Not surrounded by a membrane
Two components
Ribosomal RNA (rRNA)
Encoded by a gene
Not translated
Forms the ribosome’s “skeleton”
Proteins
Attached to the rRNA scaffolding
A Closer Look at Translation
Ribosomes
Two subunits
Each is comprised of rRNA and protein
When the subunits are joined, three binding sites exist
E, P, and A
tRNAs bind to these sites during translation
Simultaneously bind to mRNA and tRNAs during translation
A Closer Look at Translation
Steps in translation
mRNA binds to small ribosomal subunit
First tRNA binds to an AUG codon
“Start”codon
tRNA anticodon is complementary to the mRNA codon
tRNA already carries the amino acid methionine
“Loaded”
A Closer Look at Translation
Steps in translation
Large ribosomal subunit joins the ribosome
First tRNA is in “P”site
Second loaded tRNA arrives
Attaches to “A”site
Anticodon complementary to mRNA’s second codon
A Closer Look at Translation
Steps in translation
Ribosome transfers met (aa1) to aa2
Covalent linkage to aa2
Met no longer attached to its tRNA
Ribosome shifts one codon to the right
First tRNA now in “E”site
Second tRNA now in “P”site
“A”site is open
A Closer Look at Translation
Steps in translation
“Unloaded”tRNA leaves “E”(“exit”) site
Can get “reloaded”and used again
New loaded tRNA arrives
Attaches to “A”site
Anticodon complementary to mRNA’s third codon
A Closer Look at Translation
Steps in translation
Ribosome transfers dipeptide to aa3
Covalent linkage to aa3
Tripeptide formed
met-aa2-aa3-tRNA
Ribosome shifts one codon to the right
Repeat
A Closer Look at Translation
Steps in translation
Ultimately, the codon in the ribosome’s “A”site will be a “stop”codon
UAG, UGA, or UAA
A Closer Look at Translation
Steps in translation
Ribosome transfers dipeptide to aa3
Covalent linkage to aa3
Tripeptide formed
met-aa2-aa3-tRNA
Ribosome shifts one codon to the right
Repeat
A Closer Look at Translation
Steps in translation
Ultimately, the codon in the ribosome’s “A”site will be a “stop”codon
UAG, UGA, or UAA
A Closer Look at Translation
Steps in translation
No tRNAs exist that can bind to stop codons
Translation is terminated
Polypeptide chain is severed from its tRNA
Polypeptide is released
Entire translation apparatus is disassembled
Translation
Translation video
A Closer Look at Translation
Translation proceeds very quickly
E. coli can polymerize 40 amino acids per second
A second ribosome can begin translation before the first ribosome is even done
In fact, many ribosomes can simultaneously translate a single mRNA
A Closer Look at Translation
Translation proceeds very quickly
In prokaryotes, translation can even begin before transcription is complete
Why is this not true of eukaryotes?
14.5 Genetic Regulation
Not all genes are expressed in all cells at all times
To do so would be wasteful
e.g., The insulin gene is expressed only in cells of the pancreas, and not always at the same level
Gene expression is regulated
Genetic Regulation
The DNA in one of your cells is about six feel long uncoiled
Of this DNA, less than 1.2% encodes proteins
Less than one inch of the six feet
Most of our DNA is noncoding
Some of it has regulatory functions
This DNA exists both within and between genes
Genetic Regulation
When a gene is transcribed, noncoding sequences within the coding sequences are also transcribed
Exons are coding sequences
Introns are intervening “junk”sequences
These sequences must be removed before the mRNA is functional
Removal of introns is termed “splicing”
Genetic Regulation
Splicing
The initial RNA transcript contains both exons and introns
Enzymes cut the DNA at the exon/intron boundary
The intron is discarded
The exons are spliced back together
Genetic Regulation
Certain genes control the development of their mid-body (“thoracic”) structures
Hoxc8 is one of these genes
This gene is nearly identical in reptiles, birds, and mammals
Animal thoracic structures differ
A chicken has 7 vertebrae
A mouse has 13 vertebrae
Genetic Regulation
How do nearly identical genes direct these different outcomes?
The mouse Hoxc8 gene is transcribed more than the chicken Hoxc8 gene
More transcription more protein broader distribution of the protein more vertebrae
Why does this gene get transcribed more in the mouse than in the chicken?
Genetic Regulation
Transcription begins at a DNA sequence termed a “promoter”
RNA polymerase binds to the promoter
The expression of a gene is largely determined by the ability of RNA polymerase to bind to the gene’s promoter
Genetic Regulation
Enhancer elements often exist upstream of the promoter
Proteins bind to enhancer elements
This binding can make it easier for RNA polymerase to bind
Expression of the gene is increased
Genetic Regulation
The Hoxc8 enhancer sequence differs between the mouse and the chicken
The sets of proteins that bind to these enhancer elements differ between the species
Genetic Regulation
These enhancer-binding proteins have different effects in these two species
Transcription greatly enhanced in mouse
Transcription mildly enhanced in chicken
Genetic Regulation
RNA regulates DNA
Most RNA transcribed is mRNA
mRNA is translated into protein
Some RNA is not translated into protein
tRNA
rRNA
microRNAs
Genetic Regulation
All microRNAs identified to date reduce the production of specific proteins
Interfere with mRNAs
Target mRNAs for destruction
Genetic Regulation
All organisms possess genes
Eukaryotes possess thousands, though the numbers differ between species
Humans possess ~ 20,000 –25,000
Genetic Regulation
The number of genes in different eukaryotes does not vary that extensively
The regulation of these genes varies more extensively
We likely contain more regulatory DNA than protein-encoding DNA
Gene regulation accounts for much of the differences between species
The Magnitude of the Genetic Operation
Humans possess
20,000 –25,000 genes
3.2 billion base pairs
100 trillion cells
Epigenetics
Refers to all modifications to genes other than changes in the DNA sequence itself
Epigenetic modifications include addition of molecules, like methyl groups, to the DNA backbone
Adding these groups changes the appearance and structure of DNA
Alters how a gene can interact with important interpreting (transcribing) molecules in the cell's nucleus.
Epigenetics
Epigenetic modifications, or "marks," generally turn genes on or off
Allowing or preventing the gene from being used to make a protein
Mutations and bigger changes in the DNA sequence (insertions or deletions) change the sequence of the DNA and RNA
May affect the sequence of the protein as well
Imprinting
Adding methyl groups to backbone of DNA: marking
Marks both distinguish the gene copies and tell the cell which copy to use to make proteins
Maternal or paternal genes may be marked
Non-Mendelian patterns of genetics