DNA, RNA, and Protein Synthesis Flashcards
What makes up DNA?
deoxynucleotides = purine or pyrimidine base + deoxyribose (pentose) sugar + phosphate group
Which bases are purines?
Adenine (A) and Guanine (G)
Which bases are pyrimidines?
Thymine (T) and Cytosine (C)
What is the difference between purines and pyrimidines?
Purines = double ring Pyrimidines = single ring
What is a nucleoSIDE?
A nucleotide without the phosphate group (wihch can be mono, di, or tri)
Purine or pyrimidine base + deoxyribose (pentose) sugar
What is the polarity of polynucleotide strands?
5’ to 3’
What is it called when the two strands of DNA run in opposite directions?
Anti-parallel
Name all the bases in deoxynucleotide form.
Deoxyadenosine
Deoxyguanosine
What is the start codon?
AUG (coding for Methionine, Met)
What are the 3 stop codons?
UAA, UAG, UGA
What is specificity of the genetic code referring to?
The code is unambiguous: a particular codon always codes for the same amino acid (this does not change).
What is meant by universality?
The genetic code is virtually universal; specificity has been conserved from early stages of evolution w/ only slight differences in manner in which the code is translated (exception: mitochondria).
What is meant by degeneracy?
The genetic code is redundant: a given amino acid may have more than one codon coding for it, even though each codon corresponds to only one AA.
What is a point mutation?
Changing a single nt base on the mRNA chain.
What is a silent mutation?
Codon w/ the changed base ends up coding for the same AA.
What is a missense mutation?
Codon w/ changed base codes for a different AA.
What is a nonsense mutation?
Codon w/ changed base becomes a termination codon.
What is trinucleotide repeat expansion?
A mutation in which a sequence of 3 bases repeated in tandem becomes amplified, resulting in too many copies of the triplet.
What is Huntington disease caused by?
Trinucleotide repeat expansion (TRE): amplification of CAG codon leads ot insertion of many extra glutamine residues which change secondary structure causing accumulation of protein aggregates.
What is happening in both fragile X syndrome and myotonic dystrophy?
TRE occurs in an untranslated region of a gene, resulting in decrease in amount of protein produced.
Fragile X amplifies CGG, myotonic dystrophy amplifies CUG.
What is a splice site mutation?
mutations at splice sites that alter how introns are removed from pre-mRNA molecules, producing aberrant proteins.
What is a frame-shift mutation?
When one or two nt are either deleted from or added to coding region of mRNA and the reading frame is altered.
Results in product w/ radicaly different AA sequence or truncated product.
What occurs in cystic fibrosis (CF)?
Deletion of 3 nt from the coding region of a gene, resulting in the loss of phenylalanine at the 508th position in the protein encoded by that gene. Prevents normal folding of protein, which leads to destruction by the proteasome.
Presents as production of secretions in lungs and pancreas, leading to lung damage and digestive problems.
What is needed for protein translation?
- All AAs in finished product,
- mRNA to be translated
- tRNA for each amino acid
- functional ribosomes
- energy sources
- enzymes
- noncatalytic protein factors needed for initiation, elongation, and termination steps
What is an amino acid attachment site?
Each tRNA has attachment site for specific AA at its 3’ end b/c carboxyl group of AA ester links w/ 3’-hydroxyl of ribose portion of adenine (A) nucleotide in the –CCA (unpaired base tail) sequence at the 3’ end of the tRNA.
What is a tRNA called when it is covalently attached to an amino acid? What is the AA called in this situation?
Charged; activated.
What is the anticodon?
The 3 base nt sequence that pairs w/ the specific codon on the mRNA
Specifies insertion into growing peptide chain of the AA carried by that tRNA.
What does an aminoacyl-tRNA synthase do?
Attaches a specific AA to its corresponding tRNA.
Catalyze a two-step rxn resulting in covalent attachment of carboxyl group of an AA to the 3’ end of its corresponding tRNA.
a. Requires ATP → AMP + PPi (inorganic pyrophosphate)
b. Extreme specificity contributes to high fidelity of translation of genetic message.
Also can proofread/edit: remove an incorrect AA from enzyme or tRNA molecule
What are some responsibilities of the small subunit of the ribosome?
binds mRNA and is responsible for accuracy of translation by ensuring correct base-pairing between codon in mRNA and anticodon in tRNA.
How many species of rRNAs do prokaryotes and eukaryotes have?
3 and 4, respectively
How are rRNAs generated?
By action of ribonucleases on a single pre-rRNA; some bases and riboses are modified.
What are the A, P, and E sites of the ribosome?
Binding sites for tRNA molecules:
A site binds aminoacyl-tRNA as directed by the codon currently occupying the site
P site codon occupied by peptidyl-tRNA, which carries chain of AAs that has already been synthesized.
E site occupied by empty tRNA as it is about to exit the ribosome.
Where are ribosomes located in eukaryotic cells?
free in cytosol or associated with RER
What do RER ribosomes synthesize?
proteins to be exported from the cell and those incorporated into plasma, ER, or Golgi membranes or imported into lysosomes
What do cytosolic ribosomes synthesize?
proteins required in cytosol itself or destined for nucleus, mitochondria, or peroxisomes
What protein factors are required for synthesis?
Initiation, elongation, and termination (release) factors are required for peptide synthesis.
Some perform a catalytic function, whereas others stabilize synthetic machinery.
What sources of energy are required for protein synthesis?
ATP and GTP
What are the energy demands to add one amino acid to a growing polypeptide chain?
Cleavage of 4 high-energy bonds required: 2 from ATP in aminoacyl-tRNA synthetase rxn (one in removal of PPi, one in hydrolysis of PPi to inorganic phosphate by pyrophosphatase), and 2 from GTP (one for binding aminoacyl-tRNA to A site and one for translocation step).
What are isoaccepting tRNAs?
tRNAs that recognize more than one codon for a given AA.
What kind of binding occurs b/w the codon and anticodon?
Anti-parallel (mRNA codon is read 5’→3’ by an anticodon paired in flipped 3’→5’ orientation)
What is wobble?
The mechanism by which a single tRNA can recognize more than one codon for a specific amino acid.
there don’t need to be 61 tRNA species to read the 61 codons coding for AAs.
How does wobble work?
Codon-anticodon pairing follows traditional Watson-Crick rules (C pairs with G, A pairs with U) for first two bases of codon but are less stringent for the 3rd/final base, which is not as spatially defined as the other two.
Movement of this base allows nontraditional base-pairing with the 5’-base of the anticodon (the “first” base).
What components of the translation system need to be assembled in the initiation phase?
2 ribosomal subunits, mRNA to be translated, aminoacyl-tRNA specified by the first codon in the message, GTP, and initiation factors that facilitate assembly of initiation complex.
What is the Shane-Dalgarno sequence?
Purine-rich sequence of nt bases in E. coli located 6-10 bases upstream of initiating AUG codon on mRNA.
Forms complementary base pairs w/ 16S rRNA component of small ribosomal subunit’s nt sequence near 3’ end.
Gets the small ribosomal subunit in the right spot near the initiating AUG codon.
What is a 5’ cap?
The cap structure at the 5’ end of the mRNA where the small ribosomal subunit binds in eukaryotes. Subunit will move down mRNA until it encounters AUG initiator.
Scanning process requires ATP.
What does the initiator tRNA do?
Recognizes AUG facilitated by IF-2-GTP in prokaryotes and eIF-2-GTP (plus additional eIFs) in eukaryotes.
eIF = eukaryotic initiating factors
Where does the charged initiator tRNA enter the ribosome?
P site on small subunit, carrying either formylated (prokaryotes) or non formylated (eukaryotes) Methionine on the N-terminus.
Large ribosomal subunit joins complex, and functional ribosome formed w/ charged initiating tRNA in P site (A site is empty).
What is elongation?
Adding amino acids to carboxyl end of growing polypeptide chain as the ribosome moves from the 5’ end to the 3’ end of the mRNA being translated.
What is decoding?
Delivery of aminoacyl-tRNA whose codon is next on the mRNA template in the ribosomal A site.
What is the formation of the peptide bond catalyzed by?
Peptidyltransferase, an activity intrinsic to 23S rRNA found in lg ribosomal subunit.
What happens post peptide bond formation in elongation?
what was attached to P site tRNA is now linked to the amino acid on the tRNA at the A site.
What is translocation?
The ribosome advancing 3 nucleotides toward the 3’ end of the mRNA after the peptide bond is formed and the peptide chain moves from the P to the A site tRNA.
Causes movement of uncharged tRNA from Pe to E site for release and movement of peptidyl-tRNA from A to P site.
When does termination occur?
When 1 of 3 termination codons moves into the A site.
What structure recognizes the termination codons?
Release factors RF-1 (recognizes UAA and UAG) and RF-2 (UGA and UAA) in eukaryotes.
In eukaryotes, eRF = RF1, 2; eRF-3 = RF-3
How does termination proceed?
Binding RFs results in hydrolysis of bond linking peptide to tRNA at P site, causing new protein to be released from the ribosome. RF-3-GTP then causes release of RF-1 or RF-2 as GTP is hydrolyzed
What is a polysome?
Complex of one mRNA being translated by a number of ribosomes.
How can translation be regulated in eukaryotes?
Phosphorylation of eIF-2 to make it inactive.
Also, proteins that bind mRNA and inhibit its use by blocking translation or extend its use by protecting it from degradation. (this is true for prokaryotes as well)
What is protein targeting?
Addition of amino acid sequence to proteins either during or after synthesis to direct them to their final locations.
What is trimming protein modification?
Removal of portions of protein chain from large, inactive precursor molecules by specialized endoproteases resulting in the release of an active molecule.
What are the types of covalent attachment protein modifications?
Phosphorylation, glycosylation, and hydroxylation.
What is N-glycosylation?
Addition of carbohydrate chains to the amide nitrogen of asparagine (occurs in ER).
What is O-glycosylation?
Addition of carbohydrate chains built sequentially on hydroxyl groups of serine, threonine, or hydroxylysine (occurs in Golgi).
Where does hydroxylation occur?
Proline and lysine residues of alpha chains of collagen; performed by vitamin c-dependent hydroxylases in ER.
Is protein folding spontaneous or does it need help?
Both. Can be folded spontaneously as a result of primary structure or facilitated by chaperone proteins.
What happens to defective proteins or ones that have rapid turnover?
Marked for destruction by ubiquitin (small, highly conserved protein). Proteasome will rapidly degrade the protein based on this signal.
Proteasome: a macromolecular, ATP-dependent, proteolytic system located in the cytosol.
What are the protein factors involved in initiation for protein synthesis in prokaryotes and eukaryotes?
- (e)IF-2-GTP: brings charged initiating tRNA to P site
- (e)IF-3: prevents association of subunits
(IF = initiating factor; eIF = eukaryotic initiating factor)
What are the protein factors involved in elongation for protein synthesis in prokaryotes and eukaryotes?
- EF-Tu-GTP/EF-1(alpha)GTP: brings all other charged tRNAs to A site
- EF-Ts/Ef-1(beta)(gamma): guanine nucleotide exchange factor
- EF-G-GTP/EF-2-GTP: translocation
(EF = elongation factor; prokaryotic factors listed first)
What are the protein factors involved in termination for protein synthesis in prokaryotes and eukaryotes?
- RF-1 and RF-2 / eRF: recognize “stop” codons
- RF-3-GTP / eRF-3-GTP: release of other RFs
((e)RF = (eukaryotic) release factors; prokaryotic factors listed first)
What additional pairings are allowed in wobble?
G with U
I with U, C or A
