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CHAPTER 3 OUTLINE
GENE FUNCTION
TRANSCRIPTION
RNA differs from DNA.
-single-stranded
-uracil instead of thymine
-ribose instead of deoxyribose
For any gene there is a transcribed strand (template) and a non-template strand.
RNA polymerase makes RNA in a 5’ to 3’ direction, directed by a template, which is
anti-parallel to the transcript.
Initiation involved interaction of RNA polymerase with the promoter. There are
consensus target sequences in the promoter that are critical for these interactions. The
strength of the promoters is to some extent a function of how close the target sequences
are to the consensus. Mutations in these target sites can have UP or DOWN effects
depending on whether the resulting sequences are closer or farther from the consensus.
Elongation
Nucleoside triphosphates are added to the 3’ end of the growing polynucleotide chain.
The identity of the incorporated bases is dictated by the template sequence.
Termination
Termination is dependent on specific nucleotide sequence signals. A common motif in
prokaryotes is the hairpin loop structure, followed by poly-U sequence.
Unlike prokaryotes, where there is one principle RNA polymerase, transcription in
eukaryotes involves three different RNA polymerases
-RNA polymerase I (rRNAs)
-RNA polymerase II (mRNAs)
-RNA polymerase III (other small functional RNAs)
In eukaryotes, RNAs typically undergo processing after being made.
-capping at the 5’ end
-polyadenylation at the 3’ end
-splicing (removal of introns)
For splicing there are conserved sequence targets in the intron sequence
(GU_________A___AG). Splicing occurs by two transesterification events, resulting in
the excised lariat + spliced exons.
TRANSLATION
Information in the polynucleotide chain is converted to protein, a chain of amino acids.
The amino acids are joined by peptide bonds, involving condensation between carboxyl
and amino ends. There is a wide variety of amino acids that differ by virtue of their R
groups.
Levels of protein structure:
1. Primary = the linear amino acid sequence
2. Secondary = periodic structures formed by the polypeptide chains that are
stabilized by H bonds. Alpha-helix and beta-pleated sheet.
3. Tertiary = folded secondary structure
4. Quaternary = Interaction between polypeptide subunits
GENETIC CODE
The genetic code is a triplet code, which is redundant, unambiguous and universal. It
includes punctuation, such as start and stop codons.
Transfer RNA is the key intermediary in translating. It contains an anticodon at one end
(capable of interacting with mRNA codons) and an amino acid at the other end. A family
of aminoacyl tRNA synthetases ensures that the tRNAs have the appropriate amino acids.
A “Wobble” at the 3’ position of the codon permits the tRNA anticodon to recognize
more than one codon:
G in the anticodon will base pair with either C or U
U in the anticodon will base pair with either A or G
I (inosine) in the anticodon with base pair with C or U or A
The ribosome is the site of protein synthesis. It is made of one large and one small
subunit, each of which is composed of a complex of rRNA and ribosomal proteins.
MECHANICS OF TRANSLATION
Initiation
In prokaryotes, the Shine-Dalgarno sequence in the 5’ UTR pairs with the 16s rRNA in
the small subunit of the ribosome, permitting proper positioning of the start codon of the
mRNA. The start codon is AUG (or GUG) and is recognized by a specific tRNA that
carries a formylated met.
In eukaryotes, the 5’ cap interacts with the ribosome, at which point the mRNA is
scanned for the start codon (AUG). The initiator tRNA carries methionine.
The end result is that the tRNA is now located in the P site (peptidyl) along with the start
codon of the mRNA and the ribosome is poised for elongation.
Elongation
The next codon is located in the A site (aminoacyl), the point where tRNAs carrying
aminoacids gain entry. The identity of the amino acid is dictated by the codon-anticodon
interaction. A peptide bond is formed between the carboxyl end of the growing
polypeptide chain and the amino end of the incoming amino acid. The polypeptide chain
then becomes associated with the A site, before being translocation to the P site. The
result is that the A site is opened up once again for the addition of the next amino acid.
Termination
Once the stop codon is in the A site, a release factor causes chain termination and the
ribosome dissociates from the mRNA.
PROTEIN FUNCTION
Beadle and Tatum’s studies with Neurospora established clearly the relationship between
genes and their gene products, by identifying mutations that corresponded to specific
steps in biochemical pathways. The result was the one gene-one enzyme hypothesis,
which became further refined as the one gene-one polypeptide hypothesis.
Mutations and human disease
Garrod and alkaptonuria
Errors in metabolism
Mutant nomenclature
-denoting recessive vs dominant alleles
-types of mutations: point mutations, frameshifts, deletions, etc. and their gene targets.
OUTPUT in terms of function: null, leaky, silent
Haplo-sufficient vs haplo-insufficient wild type genes.
Suggested Problems for Chapter 3: 1-3, 9-12, 16, 18-19, 24-25
Make sure to look at the solved problems (pp. 86-87) as well.