Transcript
Management of Genetic Information
Learning objectives
Understand the mechanism of DNA replication, RNA synthesis and protein synthesis
Flow of genetic information
Two possible models of the DNA replication
Expt by Meselson-Stahl proved the semiconservative model of replication
Which direction does replication go? Major enzyme: DNA polymerase III
DNA double helix unwinds at a specific point called an origin of replicationorigin of replication
Polynucleotide chains are synthesized in both directions from the origin of replication; DNA replication is bidirectionalbidirectional in most organisms
At each origin of replication, there are two replication replication forksforks, points at which new polynucleotide chains are formed
There is one origin of replication and two replication forks in the circular DNA of prokaryotes
In replication of a eukaryotic chromosome, there are several origins of replication and two replication forks at each origin
Replication in prokaryotes
Replication in eukaryotes
DNA synthesis based on two template strands: leading strand and lagging strand templates; mechanism in prokaryotes is presented
DNA is synthesized from its 5’ -> 3’ end (from the 3’ -> 5’ direction of the template) the leading strandleading strand is synthesized continuously in
the 5’ -> 3’ direction toward the replication fork the lagging strandlagging strand is synthesized
semidiscontinuously (Okazaki fragments)Okazaki fragments) also in the 5’ -> 3’ direction, but away from the replication fork
lagging strand fragments are joined by the enzyme DNA ligaseDNA ligase
Replication fork
Enzymes and proteins in DNA replication
The action of DNA polymerase
Why 53’ direction?
Start of DNA replication
Unwinding DNA gyrase introduces a swivel point in
advance of the replication fork a helicase binds at the replication fork and
promotes unwinding single-stranded binding (SSB) protein protects
exposed regions of single-stranded DNA
Primase catalyzes the synthesis of RNA primer Synthesis
catalyzed by Pol III primer removed by Pol I DNA ligase seals remaining nicks
Summary of DNA replication in prokaryotes DNA synthesis is bidirectional DNA synthesis is in the 5’ -> 3’ direction
the leading strand is formed continuously the lagging strand is formed as a series of
Okazaki fragments which are later joined
DNA polymerases Five DNA polymerases have been found to exist in
E. coli Pol I is involved in synthesis and repair Pol II, IV, and V are for repair under unique conditions Pol III is primarily responsible for new synthesis
Eukaryotic DNA replication
Not as understood as prokaryotic. Due in no small part to higher level of complexity.
Cell growth and division divided into phases: M, G1, S, and G2
DNA replication occurs during the S phase
RNA synthesis
Transcription Template is DNA Major enzyme: DNA directed RNA polymerase No need for primers 5’ to 3’ direction
RNA synthesis
Requires a promoter region in the template DNA to which the RNA polymerse will bind
Promoter 40 base pairs upstream (-40) away from the start site (+1)
Three stages:initiation, elongation, termination Termination may be
rho factor dependent – rho factor terminates synthesis
or rho factor independent – formation of a stable hairpin loop
Promoter 40 base pairs upstream (-40) away from the start site (+1)
INITIATION STEP
ELONGATION STEP
TERMINATION STEP
ρ-FACTOR INDEPENDENT- FORMATION OF HAIRPIN LOOP
Eukarotic transcription have 3 classes of RNA polymerases RNA pol I transcribes large ribosomal RNA
genes RNA pol II transcribes protein encoding gene RNA pol III transcribes small RNAs
(including tRNA and 5SRNA)
Post transcriptional modification of the eukaryotic mRNA Capping – methyl guanosine attachment at the
5’ end to protect the cleavage of the RNA by exonucleases as RNA moves out of the nucleus
Addition of poly A at the 3’ end (200-250 long) helps to stabilize the mRNA structure; increases resistance to cellular nucleases
Splicing – removal of non coding sequences (introns)
Protein synthesis
Translation Based on the m-RNA sequence, genetic
code Starts from 5’ end of the transcript Occurs in the ribosomes Activation of amino acids – attachment to the
tRNA Initiation, elongation, termination
Genetic code
Triplet nucleotide – one amino acid Nonoverlapping No punctuation Degenerate Almost universal
Initiation
Initiation factors Shine-Dalgarno sequence in mRNA 30S ribosome N-formylmet
Inhibitors of protein synthesis
Postranslational modification
Protein folding –chaperones Proteolytic cleavage (zymogens) – hydrolytic
enzymes in the gut Amino acid modifications Attachment of carbohydrates Addition of prosthetic groups
Regulation of protein synthesis and gene expression 20K to 25K genes in the human genome Only a fraction of the genes are expressed at
any given time Two types of gene expression: constitutive
and inducible Inducible genes are highly regulated –
regulatory proteins, hormones and metabolites
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