Topic 3: RNA structure, transcription, and translation
Tetracycline
antibiotic that inhibits initiation by binding the 30s and blocks the A site
Streptomycin
antibiotic that inhibits initiation by binding to the smaller 30s ribosomal subunit of bacteria Can cause loss of hearing
Rifampin
antibiotic that inhibits the transcription in prokaryotes known to interfere with contraceptives
What is an amino-acyl tRNA or charged tRNA?
it is a tRNA that has the appropriate amino acid attached to it
degenerate
the reason why there are 61 codons for 20 amino acids means any nucleotide can occupy the 3rd position of an mRNA codon
What are the modifications for primary transcript to be converted into mRNA?
1. A 5' cap and 3' PolyA tail will be added to the primary transcript. The 5' end is capped with a methyl group. Folic acid (vit. B12 is required for this) At 3' end PolyA adds ~150-250 adenines The purpose of these 2 things is to protect the transcript from degradation in the cytoplasm. Act as a signal for ribosomal binding and initiation of translation 2. Introns will be removed by a process called RNA splicing
During RNA splicing how are non-coding segments of the primary transcript, introns, removed?
1. A large ribonuclear complex called a spliceosome recognizes the ends of the introns and removes it. 2. The spliceosome then joins the exons back together. • RNA polymerase and σ factor are required. • mRNA is produced by RNA polymerase II and transcription factors. 3. This modified RNA strand is now mRNA. mRNA can now leave the nucleus and enter the cytoplasm for translation
How is RNA different from DNA?
1. RNA uses ribose as its sugar (2'C has -OH) 2. RNA uses uracil instead of thymine. Base pair is A=U 3. RNA is often single stranded
Explain the process of transcription in eukaryotes?
1. The promoter region contains an A/T rich area called the TATA box which is recognized by transcription factors 2. RNA pol II recognizes the transcription factors and binds to the DNA 3. RNA polymerase II unwinds the DNA and begins to add nucleotides complementary to the template DNA 4. the new strand of RNA separates from the template DNA and exits RNA polymerase II 5. as the polymerase reads the DNA, the DNA is rewound into a helix 6. when the termination sequence is reached on the DNA, transcription is terminated and the primary transcript is released Before the primary transcript is ready to leave the nucleus it must be processed or modified to produce mRNA
What are the major types of RNA that participate directly in protein synthesis?
1. primary RNA (not a major type but a precursor to mRNA 2. messenger RNA (mRNA) 3. transfer RNA (tRNA) 4. ribosomal RNA (rRNA) 5. small interfering RNA (siRNA) or micro RNA (miRNA)
What is required for transcription in a eukaryote?
3 RNA polymerases in eukaryotes -RNA pol I=rRNA -RNA pol II=mRNA* -RNA pol III=mRNA these polymerases use a variety of proteins called transcription factors (TF) to identify promoter regions
What are the subunits for the 70S prokaryotic ribosome?
50S and 30S
What are the subunits for the 80S eukaryotic ribosome?
60s and 40s
Why is RNA polymerase in prokaryotes a primary target for antibiotics?
Because RNA polymerase is responsible for generating all types of RNA transcripts (primary transcript, mRNA, rRNA, and tRNA)
Explain initiation?
a small ribosomal subunit attaches to the 5' methyl cap The ribosome then slides to the translation initiation site called the start codon, AUG
To produce mRNA in a eukaryote what has to happen to the primary transcript?
addition of 5' cap addition of a 3' polyadenylated tail (PolyA tail) removal of introns and rejoining of the exons (splicing) When this is done the primary transcript is said to have undergone post transcriptional modification and is now mRNA
erythromycin
antibiotic that inhibits elongation by binding the 50s ribosomal subunit and inhibiting translocation
Cloramphenicol
antibiotic that inhibits elongation by binding the 50s ribosome and blocking transfer of the amino acid to growing peptide chain
Why is the difference in eukaryotic ribosomes based by species important?
because some antibiotics specifically target prokaryotic ribosomes
Why are transcription and translation not physically separate in a prokaryote
because they lack a nucleus
chloramphenicol
binds the 50S ribosome and blocks transfer of the amino acid to growing peptide chain
How is RNA polymerase similar to DNA polymerase?
can catalyze the formation of phosphodiester bonds. The bonds form between newly synthesized RNA, and the incoming nucleotides that base pair with the DNA template reads the DNA template in the 3' to 5' direction synthesizes the new RNA strand in the 5' to 3' direction
What is mRNA?
contains the nucleotide sequence specifying the amino acid sequence of a protein has been process introns have been removed and 5' cap and poly A tail have been added
What is the difference between degenerate code and wobble pairing?
degenerate: refers to the sequence on the mRNA encoding for an amino acid, even when following strict base pairing rules wobble pairing: is the type of pairing between the tRNA with the mRNA
What is the primary transcript? What does it contain? What will it do?
directly produced by transcription contains introns will undergo further processing to become mRNA
Explain termination
elongation continues until a stop codon is encountered in the A site A protein called the release factor binds the stop codon in the A site and causes the completed polypeptide to be released from the tRNA Once the polypeptide is released all the subunits dissociate and translation is terminated
How many phases are in translation and how does it differ in prokaryotes and eukaryotes?
initiation, elongation, termination differ in the ribosomal subunits used to translate
What is rRNA?
part of the ribosome that aligns the mRNA with the tRNA for translation
How is RNA similar to DNA?
phosphate, sugar, nitrogenous base
What makes up a ribosome?
rRNA and proteins
How is RNA polymerase different from DNA polymerase?
recognizes promoters at the beginning of genes (not origins) can initiate RNA synthesis without a primer uses ribonucleotides contain 2' OH groups uses UTP in place of TTP lacks 3' to 5' exonuclease activity (proofreading) must copy the correct strand of the DNA to yield the right transcript for translation
What is tRNA responsible for? is the structure of tRNA?
responsible for recognizing the mRNA and carrying the appropriate amino acid necessary for protein synthesis has a cloverleaf shape carries amino acids bound to its 3' end to the ribosomes for protein synthesis has an anticodon which base-pairs with the codons (three nucleotides) on the mRNA
codons
sequence of 3 nucleotide bases, triplets, found in mRNA that code for specific amino acids 4 different ribonucleotides (A,C,G, and U) that could be arranged into triplets 64 codon sequences that can be generated 3 stop codons: UAA, UAG, UGA
What is siRNA and miRNA?
siRNA: small interfering which were originally derived from viral genomes miRNA micro RNA originate from the host cell genome they function in a very similar manner 17-21 nucleotides responsible for binding to mRNA and interfering with translation=RNA interference
What is the single transcription factor like protein in prokaryotes?
sigma
transcription
synthesis of RNA from a DNA template (gene) first step of protein production
Wobble hypothesis
the 5' end of the tRNA anti-codon doesn't have to follow strict base pairing rules with the 3' position codon on the mRNA So the 3rd position doesn't have to follow Watson-Crick pairing G can go with C and U
translation
the conversion of genetic information from RNA to amino acids to form polypeptides (proteins)
reading frame
the start of a reading from is where the ribosome begins to read the mRNA
Why is RNA important?
transcription, translation, translational regulation
Where does transcription and translation take place in a eukaryotic cell?
transcription: nucleus (specifically nucleolus) translation: cytoplasm only 1 protein product is produced from each transcript (most commonly)