Protein Synthesis And Mutation Flashcards
What is a gene ?
Specific sequence of nucleotides along a DNA molecule that codes for a particular polypeptide chain or RNA molecule
What does The Central dogma of molecular biology describe ?
The unidirectional flow of genetic information from DNA through RNA to polypeptide
Where does DNA replication occur in eukaryotic cell ?
Nucleus
Where does DNA replication occur in prok cell ?
Nucleoid
Where does transcription occur in euk cell ?
Nucleus
Where does transcription occur in prok cell ?
Nucleoid
Where does translation occur in euk cell ?
Cytoplasm
Where does translation occur in prok cell ?
Cytoplasm
What is RNA?
A polymer of ribonucleotides joined by phosphodiester bond
What is the pentose sugar present in RNA ?
Ribose sugar
What are the nitrogenous bases of RNA ?
Adenine (A)
Guanine (G)
Cytosine (C)
Uracil (U)
Which base does uracil replace from DNA?
Thymine (T)
Are RNA single stranded or double stranded ?
Most are single stranded but an RNA molecule may contain regions which loop back upon itself due to complementary base pairing
What are the three types of RNA involved in protein synthesis ?
Messenger RNA
Ribosomal RNA
Transfer RNA
What is mRNA ?
A single-stranded polynucleotide which carries the information of DNA out from the nucleus to ribosomes in the cytoplasm where proteins are made
What makes up a codon ?
3 mRNA bases in a row
What does a codon code for ?
A certain AA
Lifespan of mRNA ?
Short-lived (degraded once it has been translated)
What is the structure of rRNA ?
Single-stranded but may fold into different shapes
Where is rRNA synthesised ?
In the nucleolus region of the nucleus
Which RNA is a major component of ribosomes ?
rRNA
What is the characteristic shape of tRNA ?
Clover-leaf shape
How does tRNA forms its characteristic clover-leaf shape ?
It folds upon itself through complementary base pairing
What does the 5’ end of tRNA always end in ?
G (guanine)
What does the 3’ end of tRNA always end in ?
CCA
Where is the amino acid binding site located on the tRNA ?
The 3’ end
What is an anticodon ?
Specific 3 bases on tRNA that is complementary to a specific codon on the mRNA
How many different tRNAs are there ?
About 45, each specific to one AA
What is the main function of tRNA ?
To bring the appropriate AA to the ribosome for protein synthesis
What is transcription ?
Process by which a sequence of bases in the DNA template of a gene is copied onto a complementary sequence of bases in mRNA
What is a transcription unit ?
Stretch of DNA template which includes the gene that is going to be transcribed, together with its promoter and terminator
What is a promoter ?
DNA sequence that carry the information to regulate transcription
What is a terminator ?
DNA sequence that carry the information to end transcription
Are the promoter and terminator sequences coding or non-coding ?
Non-coding (they do not make up any part of the polypeptide)
How many of the 2 DNA strands are used as template for transcription ?
1
What is the DNA strand that is transcribed called ?
TAN :
Template
Antisense
Non-coding
What is the strand that is not transcribed called ?
Non-template
Sense
Coding
What does RNA polymerase do ?
Catalyses formation of phosphodiester bond between neighbouring RNA nucleotides to form polynucleotide chain (mRNA)
How does RNA polymerase recognise the DNA template ?
By the presence of promoter
What are the three stages of transcription ?
- Initiation
- Elongation
- Termination
What binds to TATA box at promoter in transcription in euk ?
Collection of proteins called the general/basal transcription factors recognise and bind to the TATA box at the promoter
What do the transcriptional factors in euk do ?
Recruit the RNA polymerase and ensure its correct positioning onto the promoter
What makes up the transition initiation complex (TIC) in transcription in euk ?
RNA polymerase and the general transcription factors bound at the promoter
What does RNA polymerase do in transcription initiation in euk?
Unwind the DNA double helix in front of it by breaking the H bonds between the complementary bases to expose the nucleotides
What is the region of DNA undergoing transcription in euk called ?
Transcription bubble
Which direction does RNA polymerase move in (euk transcription) ?
3’ to 5’ direction with respect to the template strand
The newly synthesised RNA strand is elongated in what direction (euk transcription) ?
5’ to 3’ direction with respect to growing RNA chain
(euk transcription)
Where does the energy for the formation of phosphodiester bonds between the ribonucleotides come from ?
Removal of the two phosphate groups from the incoming ribonucleoside triphosphates
Does DNA remain unwounded after transcription ?
No it rewinds once RNA polymerase moves on
(euk transcription)
What happens to RNA transcript when DNA rewinds after transcription is done ?
It peels away from the DNA
Does RNA polymerase require free 3’ -OH fro synthesis of new strand ?
No
What does the terminator sequence code for ?
Polyadenylation signal (AAUAAA)
(euk transcription)
What happens when the terminator sequence is transcribed ?
Certain proteins are recruited to cleave the mRNA transcript at a point downstream of the polyadenylation signal thus halting further elongation
What happens to the RNA polymerase on the DNA strand after transcription is completed ?
It dissociates
(Prok transcription)
What competent recognises the promoter ?
The sigma factor which is a part of the RNA polymerase
When is the sigma factor released ?
Once the RNA polymerase is bound to the DNA at the promoter, unwinds the DNA double helix and synthesises about 10 nucleotides of mRNA, the sigma factor is then released
What does the release of sigma factor allow ?
Allows the RNA polymerase to move forward and continue transcription
What does the terminator sequence in prok transcription code for ?
RNA which forms a hairpin loop resulting in termination of transcription as the RNA dissociates from the template
Is the process of transcription elongation in euk the same as that in prok ?
Same
What are the differences between transcription initiation in euk and that in prok ?
Factor that recognises and bind to promoter - in euk is the general transcription factors, in prok is the sigma factor of the RNA polymerase
Formation of transcription initiation complex - in euk TIC formed with general transcription factors and RNA polymerase, in prok no TIC formed
What is the difference between transcription termination in euk and that in prok ?
Terminator sequence codes for - in euk is polyadenylation signal, in prok is for RNA which forms hairpin loop causing RNA to dissociate from template strand
In euk protein is required to cleave mRNA after polyadenylation signal, in prok mRNA dissociates on its own after transcription of termination sequence
What occurs to the pre-mRNA in euk ?
Post-transcriptional modification to form mature mRNA
Where does post-transcriptional modification occur in a cell (euk) ?
Nucleus
What are te three types of post-transcriptional modifications ?
5’ capping
3’ capping
Splicing
What is 5’ capping ?
Addition of 7-methylguanosine residue to the 5’-terminal end of the pre-mRNA transcript catalysed by mRNA guanyltranferase
What is the enzyme catalysing addition of 5’ cap ?
mRNA guanyltranferase
What are the functions of the 5’ cap ?
- Forms barrier to protect mRNA from degradation by 5’ exonucleases
- Helps in export of mature mRNA from nucleus to cytoplasm
- Helps mRNA bind to ribosomes for translation
What are introns ?
DNA region within the gene that is not translated into proteins
What are exons ?
DNA region within the gene that is translated into proteins
What is splicing ?
Process where introns are excised (removed) and exons are spliced together to produce mature mRNA
Where does splicing take place ?
Nucleus
What are splice sites ?
The start and end of an intron sequence on the pre-mRNA, tend to start with ‘GU-‘ and end with ‘-AG’
How are spliceosomes formed ?
When several snRNP bind to the splice sites and complex with each other, forming spliceosome
What are snRNP ?
Small nuclear ribonucleoproteins which are protein complexed with snRNA
How does formation of spliceosome allow for splicing to occur ?
Formation of spliceosome brings the exons upstream and downstream of an intron close together, folding the pre-mRNA into specific 3D conformation for splicing
The spliceosome then excises the introns and splices the exons together
What is alternative splicing ?
Conversion of one type of pre-mRNA into more than one type of mature mRNA by retaining and splicing only certain exons
Why are exons and introns important ?
Allows for a single gene to encode more than one type of polypeptide
Why is alternative splicing important ?
Allows one gene to give rise to more than one polypeptide thus fewer genes are needed
Are all introns removed from pre-mRNA in alternative splicing ?
Yes
What is 3’ polyadenylation ?
Addition of poly-A tail to 3’ end of pre-mRNA catalysed by poly(A) polymerase
What enzyme is involved in addition of 3’ poly-A tail?
Poly(A) polymerase
What are the functions of the 3’ poly-A tail ?
- Protect the mRNA fro degradation by 3’ exonucleases
- Facilitate transport of mRNA out of nucleus into cytoplasm
- Stabilised mRNA as a template for translation
What are the four features of the genetic code ?
- Triplet code
- Universal
- Degenerate but not ambiguous
- Non-overlapping
What is an mRNA codon made up of ?
3 consecutive nucleotide bases
There are only __ ribonucleotide bases to specify __ AAs
Only 4 bases to specify 20 AAs
Why 3 nucleotide bases as 1 codon ?
Smallest possible number to be able to give rise to the 20 AAs
- is each codon only 2 bases, there would only be 16 AAs
What are the nucleotide bases for the three stop codons ?
UAG
UGA
UAA
What is the nucleotide base of the start codon ?
AUG
What does the start codon code for in eukaryotes ?
Methionine
What do the stop codons code for ?
Do not code for AAs
How is the genetic code universal ?
The same triplet of bases code for the same AAs in all organisms
What does having a degenerate genetic code mean ?
Means that more than one codon can code for the same AA
What does genetic code being not ambiguous mean ?
Means that each codon only codes for one AA
What does it mean when the genetic code is non-overlapping ?
Start codon AUG defines the start of mRNA reading frame, the mRNA is then read sequentially one codon at a time in 5’ to 3’ direction
What is translation ?
Synthesis of a polypeptide which occurs under the direction of mRNA
Where does translation occur ?
At ribosomes in the cytoplasm
What are the four stages of translation ?
- Preparation
- Initiation
- Elongation
- Termination
How is aminoacyl-tRNA formed ?
Attachment of AA to tRNA catalyses by aminoacyl-tRNA synthetase, energy from hydrolysis of ATP is required
Are aminoacyl-tRNAs specific ?
Yes - there is one aminoacyl-tRNA specific for each AA
Where is AA bound to tRNA ? why ?
At the AA binding site at the 3’ end of tRNA
Enables AA to form a peptide bond with carboxyl end of growing polypeptide chain
What happens in translation preparation of euk ?
Formation of aminoacyl-tRNA ( amino acid activation )
Which type of ribosome are involved in translation in euk ? Subunits ?
80s ribosomes
Small 40s subunit and large 60s subunit
Which ribosomal subunit contains mRNA binding site for translation in euk ?
Small subunit
(Translation in euk)
What does the mRNA binding site in small subunit bind to ?
5’ untranslated region (UTR) of mRNA
What are the three binding sites present in large ribosomal subunit ?
Aminoacyl-tRNA site : A site
Peptidyl-tRNA site : P site
Exit site : E site
What does the A site of large ribosomal subunit in translation hold ?
Holds the tRNA carrying the next AA to be added to the growing polypeptide
What does the P site of the large ribosomal subunit in translation hold ?
Holds the tRNA carrying the growing polypeptide
What does the E site of the large ribosomal subunit in translation hold ?
The free tRNA before it leaves the ribosome
(Translation in euk)
What AA does the initiator tRNA carry ?
Methionine
Where does the initiation tRNA bind to during translation initiation in euk ?
Small ribosomal 40s subunit
(Translation in euk)
What are initiation factors ?
Proteins that bind to small subunit after initiator tRNA has bound, to stabilise the structure
When does small ribosomal 40s subunit bind to mRNA ?
After initiator tRNA has already bound
How does small ribosomal 40s subunit bind to mRNA ?
Via mRNA binding site on subunit which contains rRNA complementary to 5’ UTR of mRNA
What happens after small ribosomal 40s subunit binds to mRNA ?
It moves downstream along mRNA until it reaches start codon (AUG) and anticodon of initiator tRNA forms complementary base pair with it
What is the translation initial complex made of in euk ?
Small ribosomal 40s subunit bound to large ribosomal 60s subunit
When translation initiation in euk is completed, what occupies which sites on the large ribosomal 60s subunit ?
Initiator tRNA occupies P site
A site is vacant, ready for the next aminoacyl-tRNA
What occurs in translation initiation in euk (quick overview) ?
- binding of initiator tRNA to small 40s subunit
- binding of small 40s subunit to mRNA
- scanning downstream of small 40s subunit for AUG
- formation of translation initiation complex
What are the three stages of translation elongation in euk ?
- Aminoacyl-tRNA binding
- Peptide bond formation
- Translocation
What happens in aminoacyl-tRNA binding, the first step of translation elongation cycle in euk ?
Corresponding aminoacyl-tRNA for the next codon binds to mRNA at A site of ribosome via complementary base pairing
What enzyme catalyses formation of peptide bond during translation in euk ?
Peptidyl transferase
Where is peptidyl transferase found in euk ?
It is part of the large 60s ribosomal subunit
What occurs in peptide bond formation, second step of translation elongation cycle in euk ?
Peptide bond formed between AA/peptide on tRNA at P site and AA on tRNA at A site, catalyses by peptidyl transferase
What occurs in translocation, last step of translation elongation cycle in euk ?
Ribosome shifts 3 nucleotides down mRNA in 5’ to 3’ direction wrt mRNA
Free tRNA in P site shifts to E site to be released
Peptidyl-tRNA in A site is shifted to P site
A site is now empty and aligned to next codon and new aminoacyl-tRNA enters
Is energy needed for translocation process of ribosome in euk during translation elongation ?
Yes - energy from the hydrolysis of GTP provides energy for the translocation process
What is accepted in A site when stop codon is reached ?
Release factor instead of aminoacyl-tRNA
(Translation in euk)
What does the release factor result in ?
Addition of water molecule instead of AA to teh polypeptide chain, causing hydrolysis of polypeptide from tRNA in P site, releasing polypeptide through tunnel in large subunit
Is translation preparation in proks the same as in euk ?
Yes
What ribosome are involved in translation in prok ? Subunit ?
70s ribosomes - small 30s subunit and large 50s subunit
What does mRNA binding site on small 30s subunit in proks bind to ?
Shine-dalgarno sequence near 5’ end of mRNA
What are the 3 binding sites present in large 50s subunit ?
Aminoacyl-tRNA site : A site
Peptidyl-tRNA site : P site
Exit site : E site
How does small 30s subunit bind to mRNA in prok translation ?
Through rRNA at mRNA binding site of small subunit which is complementary to the shine-dalgarno sequence
Is there need for small 30s subunit in prok to scan down mRNA to find AUG like in euk ? Why ?
No
When mRNA binds to small ribosomal submit, AUG is already positioned at P site
What AA does initiator tRNA in prok carry?
Formal-methionine
Does translation initiation complex form in translation in prok ?
Yes
What occurs in translation initiation in prok (short overview) ?
- small subunit bind to mRNA
- initiator tRNA bind to AUG at P site
- large subunit bind to small subunit to form TIC
Is translation elongation in prok the same in euk ?
Yes
Is translation termination in prok the same in euk ?
Yes
What are the three differences between translation in euk and prok ?
AA attached to initiator tRNA - methionine in euk, formyl-methionine in prok
Type of ribosome - 80s in euk, 70s in prok
mRNA sequence in translation initiation - 5’ UTR in euk, Shine-Dalgarno in prok
What are polyribosomes ?
Structure formed when multiple ribosomes are translating an mRNA simultaneously
Are polyribosomes found in both prok and euk ?
Yes
What is the function of a polyribosome ?
Allow cell to make copies of the polypeptide very quickly
Can transcription and translation occur simultaneously ?
Yes in prok - lack of nuclear envelop
No in euk
How do polypeptides synthesised by ribosomes on rER enter cisternas ?
Through use of signal peptide attached to the N-terminus of proteins allowing the polypeptides to enter through channel proteins on the membrane of rER
Where are the proteins synthesised by free ribosomes headed?
Generally remain in cytoplasm or transported into various organelles for cellular function
Where are the proteins synthesised by ribosomes on rER headed ?
Generally secreted out of cell, inserted into various membranes as membrane proteins or are transported into the lysosome
What is a gene mutation ?
Change in DNA sequence
What could gene mutation result in ?
Change in sequence of nucleotides transmitted to mRNA, resulting in different AA being coded, forming abnormal polypeptide due to change in 3D conformation, affecting the phenotype of organism or resulting in heritable diseases
What are the two general types of point mutations ?
Addition and deletion
Substitution
When does base pair addition and deletion occur ?
When one or more base pairs are either add or deleted from a gene
What does addition and deletion of base pairs result in ?
Frameshift mutation where reading frame of mRNA after site of addition or deletion is changed, causing the primary structural of resultant polypeptide to be changed drastically
What kind of protein does frameshift mutation result in ?
Non-functional protein
When does base pair substitution occur ?
When one base pair is replaces with another pair of nucleotides
When does base pair substitution result in significant change in protein ?
Missense mutation
Nonsense mutation
What is a missense mutation ?
When substitution lead to change in codon coding for different AA resulting in change in primary structure of protein
What is a nonsense mutation ?
When substitution leads to formation of stop codon resulting in premature termination of translation and production of truncated and often non-functional protein
When does base pair substitution result in insignificant changes in the protein ?
When the new AA from the missense mutation has similar properties as the one it replaces or when new AA is in a part of protein which the exact sequence of AA is not essential to its activity
When does base pair substitution result in no change in AA ?
Silent mutation (degenerate code)
What is a silent mutation ?
When original codon in changed into another codon coding for same AA
What is chromosomal aberration ?
Change in structure of chromosome or change in number of chromosomes in a cell
When does structural aberration occur ?
Prophase I of meiosis
What are the four types of structural aberration ?
Translocation
Duplication
Inversion
Deletion
(Structural aberration)
What is translocation ?
Portion of a chromosome detached and re-joins at a different point on the same chromosome or different chromosome
(Structural aberration)
What is duplication ?
Portion of a chromosome replicates and set of genes become repeated
(Structural aberration)
What is inversion?
Portion of chromosome detaches and re-joins in an inverted manner (overall genotype may be unchanged but phenotype may be altered)
(Structural aberration)
What is deletion ?
Loss of portion of chromosome - usually lethal as genes are removed
What are the two types of numerical aberrations ?
Aneuploidy
Polyploidy
What is aneuploidy ?
Condition in which there is a missing or extra chromosome resulting in monosomy or trisomy for a particular chromosome
What is trisomy 21 ?
Condition where cells have an extra chromosome 21 resulting in Down syndrome
What is polyploidy ?
Condition where there are more than 2 sets of chromosomes resulting in triploid or tetraploid
What are the two types of polyploidy ?
Autopolyploid
Allopolyploid
What is autopolyploid ?
When all chromosomes are derived form the same species or when a basic set of chromosomes is multiplied
What is allopolyploid ?
When the chromosomes are derived from 2 different species (usually in plants)
What is sickle cell anemia ?
A homozygous recessive disorder
What if an individual only has 1 mutated allele, HbS ?
Suffer from sickle cell trait
What mutation results in sickle cell anemia ?
Single base pair substitution in gene coding for ß-globin chain in which T is replaced with A in DNA template strand
What change occurs in codon for mutated gene coding for HbS ?
Codon changed from GAG coding for glutamic acid to GUG coding for valine
What is the significant of valine being coded for instead ?
Valine is hydrophobic while glutamic acid is hydrophilic thus decreasing the solubility of deoxygenated HbS
What happens to sickle cells when oxygen level is low ?
HbS molecules precipitate out of solution to form rigid fibres causing RBC to change from circular biconcave shape to sickle shape
What are the effect of sickle cell anemia ?
- HbS molecule is less efficient in carrying oxygen compared to normal HbA
- sickled RBCs are more fragile than normal RBCs, they haemolyse (break down) readily resulting in anemia
- sickled RBCs may obstruct blood vessels and interfere with circulation, depriving multiple organs of oxygen resulting in damage
- sickled RBCs accumulate in spleen for destruction leading to enlargement of spleen and possibly death
What results in trisomy 21?
Failure of separation of chromosome 21 during meiosis anaphase I or II
Fusion of this gamete with normal gamete results in extra copy of chromosome 21
What are the effects of Down syndrome ?
- characteristic facial features and short stature
- mental retardation
- below average life-span
- most are sexually undeveloped and sterile
