Lecture 2: Central Dogma Flashcards
What is the sequence for the flow of information?
DNA -> RNA -> Protein
DNA replication is semidiscontinuous. What does this mean?
•The leading strand copies continuously
•The lagging strand copies in segments (Okazaki fragments) which must be joined
DNA Polymerase I
•abundant but not ideal for replication
•Rate (600 nucleotides/min) is slower than observed for replication fork movement
•Has low processivity
•Its primary function is in clean-up
DNA Polymerase III
The principal replication polymerase
DNA Polymerase II, IV, V
Are involved in DNA repair
DNA Pol a
•Used to make primers for Okazaki fragments
•Has primase but no 3’ -> 5’ proofreading
DNA Pol e
•Used in leading strand synthesis
DNA pol g
•Used in lagging strand synthesis
•Comparable to bacterial DNA Pol III
•Has 3’ -> 5’ proofreading
What are the final steps in the Synthesis of the Lagging Strand?
•DNA ligands makes a bond between a 3’-OH and a 5’-PO4
•5’-PO4 must be activated by attachment of AMP
•3’-OH nucleophile attacks this phosphate, displacing AMP
Uracil
Basic site
8-Oxoguanine
Single-Strand break
•Base-Excision repair (BER)
(6-4)
Bulky adduct
CPD
•Nucleotide-excision repair (NER)
Interstate cross-link
Double-strand break
•Recombinational repair (HR, EJ)
A-G Mismatch
T-C Mismatch
Insertion
Deletion
Mismatch repair
A-G Mismatch
T-C Mismatch
Insertion
Deletion
Mismatch repair
What is the end replication problem?
•Leading strand is synthesized to the end of the chromosome
•Lagging strand utilizes RNA primers which are removed
•The lagging strand is shortened at each cell division
What is the solution to the End Replication problem?
•Telomeres
Messenger RNAs
•Encode the amino acid sequences of all the polypeptides found in the cell
Transfer RNAs
•Match specific amino acids to triplet codons in mRNA during protein synthesis
Ribosomal RNAs
•Are the constituents and catalytic appropriate amino acids
MicroRNA
•appears to regulate the expression of genes, possibly via binding to specific nucleotide sequences
What are the E. coli Promoters?
•Consensus sequence
•rrn B P1
•trp
•lac
•recA
•araBAD
RNA Polymerase I
•synthesizes pre-ribosomal RNA (precursor for 28S, 18S, and 5.8 rRNAs)
RNA Polymerase II
•is responsible for synthesis of mRNA
•very fast (500-1000 nucleotides/sec)
•Specifically inhibited by mushroom toxin a-amanitin
•Can recognize thousands of promoters
RNA Polymerase III
•Makes tRNAs and some small RNA products
What RNA Polymerase does Plants have?
•Plants appear to have RNA polymerase IV that is responsible for the synthesis of small interfering RNAs
What has their own RNA polymerase?
Mitochondria
Promoters recognized by Eukaryotic RNA Polymerase II
•Various regulatory sequences
•TATA box
•lnr
Transcription at RNA II Promoters
- Pol II is recruited to the DNA by transcription factors
- The transcription bubble forms
- The CTF is phosphorylation during initiation. The polymerase escapes the promoter
- Transcription elongation is aided by elongation factors after TFIIE and TFIIH dissociate
- Elongation factors dissociate. The CTD is dephosphorylated as transcription terminates, a process facilitated by termination factors
Maturation of mRNA in Eukaryotes
- Transcription and 5’ capping
- Completion of Primary transcript
- Cleavage, Polyadenylation, and splicing
What is the function of tRNA in translation?
•Deliver the amino acids corresponding to each codon within the mRNA sequence.
How many genetically encoded amino acids are there?
•20
Four letter code in groups of two
•insufficient (16)
Four-letter code in groups of three
•Sufficient (64)
What direction is the genetic code written?
5’->3’ direction
First Codon establishes
The reading frame
If reading frame is thrown off by a base or two, what happens?
All subsequent codons are out of order
Initiation codon
AUG
Termination Codon
•UAA
•UGA
•UAG
Stages of Protein synthesis
- Activation of amino acids
- initiation of translation
- Elongation
- Termination and Ribosome recycling
- Folding and Post-translational processing
Activation of amino acids
•tRNA is aminoacylated
Initiation of translation
•mRNA and aminoacylated tRNA bind to ribosome
Elongation
•Cycles of aminoacyl-tRNA binding and peptide bond formation….until a STOP codon is reached
Termination and ribosome recycling
•mRNA and protein dissociate, ribosome recycled
Folding and post-translational processing
•Catalyzed by a variety of enzymes
Termination initiation and Elongation in E. Coli
- RNA polymerase core bonds to the DNA promoter
- Transcription bubble forms
- Transcription is initiated
- Promoter clearance is followed by elongation
- Elongation continues. S70 dissociates, and is replaced by NusA
- Transcription is terminated. NusA dissociates, and the RNA polymerase is recycled
Three rules of DNA Replication
- DNA replication is semiconservative
- DNA replication is bidirectional
- DNA replication is semicontinuous
Semiconservative
Each strand serves as a template for replication. The 2 new strands are composed of one new strand and its template (old strand)
Bidirectional
Replication occurs in both directions from a starting point, sometimes called the origin
Three stages of DNA Replication
- Initiation
- Elongation
- Termination
