Download CHAPTER 4 Notes, Part 2: FLOW OF GENETIC INFORMATION Fall

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CHAPTER 4 Notes, Part 2: FLOW OF GENETIC INFORMATION
LECTURE TOPICS: FLOW OF GENETIC INFORMATION
1)
RNA properties in general
2)
Transcription: RNA Polymerase
3)
mRNA properties
4)
Genetic Code
5)
Eucaryotic genes: structural organization
!
Accuracy [Absolute and relative]
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Signals [Starts and stops]
!
Stages [Initiation, elongation and termination]
Fall 2010
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RNA properties in general: Polynucleotides consisting of four bases (A, G, C, U),
ribose phosphate; nucleosides joined by 3' to 5' phosphodiester bonds (like DNA).
!
Base-pairing is significant in most RNA
molecules. RNA is usually single-stranded,
but it can form base-paired hairpin loops
(reversing 5' 3' orientation) with itself. A pairs
with U and G pairs with C; base paired
structures where the U of RNA pairs with A of
DNA and G-C pairs also occur [Fig. 5.19]
!
Cells contain 3 major types of RNA called mRNA (5%), tRNA (15%), and
rRNA (80%) [Table 5.2].
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Structure of RNA molecules: [ IMPORTANT!! ]
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Usually RNA is “single-stranded”
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But, the RNA forms base-paired DOUBLE-STRANDED structures by
folding back on themselves (reversing polarity) and forming base paired
structures - much like seen in DNA. Formation of these structures
depends on locations of complementary bases of the reversed polarity
sections of the RNA molecule (Figs. 5.19 and 5.20).
[Fig.5.19]
[Fig.5.20]
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Transcription: RNA Polymerase
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mRNA, rRNA and tRNA are complementary to genomic DNA (shown by DNA-RNA
hybridization experiments). (Fig.5-26)
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RNA polymerase makes an RNA which is complementary to the DNA template
strand.
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All cellular RNA is synthesized (“transcribed” from DNA) by
DNA-dependent RNA polymerase enzymes which require:
<
A template DNA, preferably double-stranded
<
Activated precursors (all 4 rNTPs-ATP, GTP, CTP, UTP
<
Mg++ in vivo
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!
RNA polymerase reaction [ Fig. 5-25]: (RNA)n + rNTP ! (RNA)n+1 + PPi
[as for DNA polymerase, reaction mechanism is nucleophilic attack of free 3'-OH
of primer on the "-P of the rNTP substrate. Pyrophosphatase also drives the
reaction forward here (PPi ! 2Pi)]
!
NOTE: RNA polymerase does not require a primer.
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Start Signals for Transcription: PROMOTERS
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Transcription begins and ends at specific sites (relative to the template strand)
called promoters. Promoter base sequences are similar in many different genes.
These sequences differ between procaryotes and eucaryotes. (Fig. 5-27)
(a) Two conserved promoter sequences are 5' to the start of transcription:
1) procaryotes: -10 (TATAAT) and -35 (TTGACA)
2) eucaryotes: -25 (TATA) and -75 (CAAT)
Stop Signals for Transcription:
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TERMINATORS
Transcription ends at sequences called
terminators. Terminators have (in the mRNA) a
GC-rich region followed by a string of U's which
can form a base-paired hairpin loop. (Fig. 5.28)
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mRNA and the GENETIC CODE:
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The concept of mRNA was based on knowing that T2 bacteriophage infection is
followed rapidly by synthesis of RNA, proteins, and new virus particles.
[Experiment: Infect E. coli cells with T2 phage and add 32P-phosphate at
same time. Then observe properties of polynucleotides that incorporate
32
P-phosphate].
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Jacob and Monod predicted properties of mRNA:
<
<
<
<
<
a polynucleotide [RNA]
base composition complementary to a DNA template(see Table 5.3)
size range to reflect the variety of protein sizes [3 bases/amino acid]
transient association with ribosomes
rapid turnover (about 2 minute ½ life in E. coli).
THE GENETIC CODE AND PROTEIN SYNTHESIS:
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Transfer RNA (tRNA) is the "adaptor" molecule which brings amino acids to
mRNA for translation of the codons into unique polypeptides.
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tRNA has two business ends:
<
<
3'-CCA-OH which accepts specific amino acids (Figs.5.30, 5.31)
anticodon which base pairs with specific codon (Fig.5.31)
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!
A codon (3 bases) specifies an amino acid.
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The genetic code is sequential, nonoverlapping, and not punctuated. [but a
few exceptions!].
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The code is degenerate (more than one codon per amino acid)
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!
Some codons - start (Fig.5.32) and stop (Table 5.4) signals. START:
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The genetic code is nearly universal. An exception, for instance, is use of
UGA (a universal stop codon) as a code word for tryptophan in human
mitochondria.
THE GENETIC CODE: SUMMARY
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A codon (3 bases) specifies an amino acid.
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Sequential and nonoverlapping.
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Degenerate (more than one codon per amino acid).
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Some codons are stop and start signals.
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The code is nearly universal [Table 5.5]. (but ex: UGA = Trp in human
mitochondrial DNA)
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The sequences of genes and their encoded proteins are colinear
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Experiments with synthetic mRNAs established codon assignments
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EUCARYOTIC GENES: (Fig.5.29)
Both ends of eucaryotic mRNA’s are modified: [5' cap and 3'-poly(A) tail ]
EUCARYOTIC GENES: STRUCTURAL ORGANIZATION
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Confusing data resolved in 1977: Eucaryotic genes have discontinuous
coding sequences. (Fig.5.33; ex., Electron microscopy)
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Most have coding sequences (exons - 3 in blue, Figure below) interrupted by
non-coding (introns) sequences. (P.136, 5th Ed.)
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!
After transcription, introns are removed and the
exons are joined accurately by splicing (Fig.5.34)
at evolutionarily conserved sequences (Fig.5.35).
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Exon polarity (5' to 3') is retained after splicing.
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Exon Shuffling: Protein domains coded by exons can be rearranged to give
proteins with new functions. (Fig.5.36)
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Alternate splicing of an mRNA can give different proteins. (ex: membranebound vs. soluble antibody)
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Almost all bacterial genes lack introns. (May have had them, but were lost
while evolving to adapt to very rapid growth?)
CHAPTER 4: Study Exercise
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Draw a typical procaryotic gene and a eucaryotic gene with two exons.
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Indicate where signals for transcription and translation would be located,
relative to each other.
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For the eucaryotic gene, indicate the intron processing (splicing signals) and
their locations, relative to the other signals.