Chapter 17: Gene Expression Flashcards
Template and coding strand
The template strand provides the pattern for the sequence of nucleotides transcribed by mRNA
The non-template strand or coding strand is the DNA strand that is complementary to the template strand
- Its condons are identical to the mRNA sequence with the substitution of thymine instead of uracil
Codons are read by the translation machinery in the 5’ → 3’ direction
Stop and start codons
AUG is the sole start codon; also codes for methionine
UAA, UAG, and UGA are all stop codons
Transcription components
RNA polymerase- enzyme that pries the two strands of DNA apart and joins complementary RNA nucleotides together
Promoter- sequence of a gene where RNA polymerase attaches to in order to initiate transcription
Transcription unit- the stretch of DNA downstream from the promoter that is transcribed
Terminator- sequence that signals the end of transcription; ONLY present in prokaryotes
Transcription
In prokaryotes
Initiation
RNA polymerase binds itself directly to the promoter and begins transcription
- Untwists DNA and exposes 10−20 nucleotides at a time for pairing with RNA nucleotides
Elongation
Adds nucleotides to the 3’ end of the growing RNA molecule similar to DNA polymerase
Termination
Transcription proceeds through the terminator sequence in the DNA which causes the polymerase to detach and release the transcript which requires no further modification
Transcription
In eukaryotes
Initiation
A collection of proteins called transcription factors help guide the binding of RNA polymerase II to the promoter
- Only after transcription factors are attached to the promoter does RNA polymerase II bind to it
- Entire complex of transcription factors and RNA polymerase II bound to the promoter is called a transcription initiation complex
- A crucial promoter DNA sequence called the TATA box helps form the initiation complex
Untwists DNA and exposes 10−20 nucleotides at a time for pairing with RNA nucleotides
Elongation
Adds nucleotides to the 3’ end of the growing RNA molecule similar to DNA polymerase at a rate of about 40 nucleotides per minute
Termination
RNA polymerase II transcribes the polyadenylation signal sequence; the RNA transcript is released 10−35 nucleotides past this polyadenylation sequence (AAUAAA)
Pre-mRNA then proceeds on for further processing
RNA processing
In eukaryotes
Enzymes in the nucleus modify pre-mRNA before the genetic message is dispatched to the cytoplasm
Both ends of the primary transcript are usually altered
Usually certain interior sections of the molecule are also cut-out and the remaining parts spliced together
Alteration of mRNA ends
In eukaryotes
The 5’ end which is synthesized first receives a 5’ cap of a modified form of guanine
At the 3’ end an enzyme adds 50−250 more adenines after the polyadenylation sequence AAUAAA forming a poly-A tail
5’ cap and poly-A tail have several function:
- Facilitate the export of mature mRNA from the nucleus
- Help protect the mRNA from degradation by hydrolytic enzymes
- Help ribosomes attach to the 5’ end once it reaches the cytoplasm
RNA splicing process
Large portions of the RNA primary transcript molecules are removed and the remaining portions reconnected
The removed segements are noncoding regions called intervening sequences or introns
The other regions called exons that remain are expressed and go to be translated into proteins
The terms introns and exons are use to describe RNA sequences and the DNA sequences that specify them
RNA splicing enzyme
The removal of introns is accomplished by a large complex made up of proteins and small RNAs called spliceosomes
Spliceosomes consist of a variety of proteins and several small nuclear ribonucleoproteins (snRNPs) that recognize the splice sites
This complex binds to several short nucleotide sequences along and intron including key sequences at each end
The intron is then released and the spliceosoe joins together the two exons that flanked the intron
Ribozymes
Ribozymes are catalytic RNA molecules that function as enzymes and can splice RNA
- Not all biological catalysts are proteins
Three properties of RNA enable it to function as an enzyme:
- It can form a three-dimensional structure because of its ability to base-pair with itself; a specific structure is essential to catalytic functioning
- Some bases in RNA contain functional groups that may participate in catalysis
- RNA can hydrogen-bond with other nucleic acid molecules which adds specificity to its catalytic activity
Functional importance of introns
A single gene can encode more than one kind of polypeptide
Many genes are known to give rise to two or more dfferent polypeptides depending on which segements are treated as exons; a process known as alternative RNA splicing
Protein often have a modular architecture that consists of discrete functional and structural regions called domains that impart different properties; different exons code for different domains
Molecular components of translation
A cell translates an mRNA message into protein with the help of transfer RNA or tRNA
Function is to transfer an amino acid from the cytoplasm to a growing polypeptide in a ribosome
Each tRNA molecule enables translation of a given mRNA codon into a specific amino acid
tRNA twists into a 3-D L-shapped structure
- Its 3’ end protrudes from one end and serves as an attachment site for a specific amino acid
- The loop extending from the other end of the L includes the anticodon which is a nucleotide triplet that base pairs with a specific mRNA codon
tRNA-amino acid specificity
The correct matching of tRNA and amino acid is carried out by a family of enzymes called aminoacyl-tRNA synthetases
The active site of each type of aminoacyl-tRNA synthetase fits only a specific combination of amino acid and tRNA
There are 20 different synthetases, one for each amino acid
The synthetase catalyzes the covalent attachment of the amino acid to its tRNA in a process driven by the hydrolysis of ATP
tRNA-mRNA specificity
Some tRNAs are able to bind to multiple codons coding for the same amino acid due to flexible base pairing between the third nucleotide base of a codon
45 tRNAs bind to more than one codon
This flexible base pairing is called wobble which is why synonymous codons for a given amino acid most often differ in their third nucleotide base
Ribosomal structure
Consists of a large and a small subunit made up of ribosomal RNA or rRNA
Ribosome has one binding site for mRNA and three binding sites for tRNA:
- A site is where tRNA enters; holds the tRNA carrying the next amino acid to be added to the chain
- P site holds the tRNA carrying the growing polypeptide chain
- E site is where the empty tRNA exits from
Stages of translation and total energy usage
Just the stages
Initiation
- Uses one molecule of GTP
Elongation
- Uses two molecules of GTP
Termination
- Uses two molecules of GTP
Total energy usage is five molecules of GTP
Initiation stage of translaiton
Brings together an mRNA, a tRNA bearing the first amino acid, and the two subunits of a ribosome
AUG start signals the start of translation and establishes the reading frame
The union of mRNA, the initiator tRNA, and the small ribosomal subunit is followed by the attachment of a large ribosomal subunit completing the translation initiation complex
- Proteins called initiation factors are required to bring all these components together
- Cell also expends energy obtained by the hydrolysis of a GTP molecule to form the initiation complex
Elongation stage of translation
A polypeptide is always synthesized in one direction from the initial methionine at the amino end, called the N-terminus, to the final amino acid at the carboxyl end, called the C-terminus
Amino acids are added one by one to the C-terminus of the growing chain
Each addition involves proteins called elongation factors and occurs in three steps:
- Codon recognition; requires one molecule of GTP
- Peptide bond formation
- Translocation; requires one molecule of GTP
Energy expenditure occurs in the first and third steps
Translation proceeds along the mRNA in a 5′ → 3′ direction
Termination of translation
Elongation occurs until a stop codon in the mRNA reaches the A site
Codons UAG, UAA, and UGA do not code for amino acids but instead act as stop signals
A release factor protein binds directly to the stop codon in the A site
The release factor causes the addition of a water molecule instead of an amino acid
This reaction releases the polypeptide and the translation assembly comes apart
- Breakdown of the translation assembly requires the hydrolysis of two more GTP molecules
Post-translational modifications
During its synthesis a polypeptide chain begins to coil and fold spontaneously to form a protein with a specific shape
Post-translational modifications may be required before the protein can begin doing its particular job in the cell
Targeting polypeptides to specific locations
There are two populations of ribosomes
- Free ribosomes mostly synthesize proteins that function in the cytosol
- Bound ribosomes make proteins of the endomembrane system and proteins that are secreted from the cell
Ribosomes are identical and can switch from free to bound
Polypeptide synthesis always begins and finishes in the cytosol unless the polypeptide signals the ribosome to attach to the ER
- Polypeptides destined for the ER or for secretion are marked by a signal peptide which targets the protein to the ER
- A signal-recognition particle (SRP) binds to the signal peptide and escorts it and its ribosome to a receptor protein translocation complex built into the ER membrane
Other kinds of signal peptides are used to target polypeptides to mitochondira, chloroplasts, or other organelles that are not part of the endomembrane system
- In these cases however translation is completed in the cytosol before the polypeptide is imported into the organelle
Translation of multiple polypeptides
Multiple ribosomes can translate a single mRNA simultaneously forming a string of ribosomes called polyribosomes or polysomes
- Enable a cell to make many copies of a polypeptide very quickly
Cells can also increase the number of copies of a polypeptide by transcribing multiple mRNAs from the same gene
Small scale genetic mutations
Point mutations are chemical changes in just one base pair of a gene and can lead to a range of effects
- Sickle-cell disease results from a point mutation which codes for a valine where a glutamic acid should be
Single nucleotide-pair substitutions
The replacement of one nucleotide and its complement with another pair of nucleotides
- Silent mutations- have no effect on the amino acid produced by a codon because of redundancy in the genetic code
- Missense mutations- still code for an amino acid but it is not the correct amino acid
- Nonsense mutations- change an amino acid codon into a stop codon nearly always leading to a nonfunctional protein
Nucleotide-pair insertions or deletions
Additions or losses of nucleotide pairs in a gene
- Frameshift mutations- occur whenever the number of nucleotides inserted or deleted is not a multiple of three; all nucleotides downstream will be improperly grouped into codons and will result in extensive missense mutations unless it occurs at or near the end of the gene
New mutations
Spontaneous mutations can occur during DNA replication, recombination, or repair
Mutagens are physical or chemical agents that can cause mutations
