GENES AND PROTEIN SYNTHESISChapter 7
ONE GENE-ONE POLYPEPTIDE HYPOTHESIS
DNA contains all of our hereditary information
Genes are located in our DNA
~25,000 genes in our DNA (46 chromosomes)
Each Gene codes for a specific polypeptide
MAIN IDEA Central Dogma
Francis Crick (1956)
OVERALL PROCESS
Transcription DNA to RNA
Translation Assembly of
amino acids into polypeptide
Using RNA
DNA molecule
Gene 1
Gene 2
Gene 3
DNA strand
TRANSCRIPTION
RNA
Polypeptide
TRANSLATIONCodon
Amino acid
KEY TERMS RNA transcription
Initiation, Elongation, Termination TATA box Introns, Exons mRNA, tRNA, rRNA Translation Ribosome Codon Amino Acids Polypeptide
DNA RNA
Double stranded Single stranded Adenine pairs with Thymine Adenine pairs with Uracil Guanine pairs with Cytosine Guanine pairs with Cytosine
Deoxyribose sugar Ribose sugar
DNA TO PROTEIN Protein is made of amino acid
sequences 20 amino acids How does DNA code for amino acid?
GENETIC CODE Codon
Three letter code 5’ to 3’ order Start codon Stop codon
AA are represented by more than one codon
61 codons that specify AA
AMINO ACIDS Abbreviated
Three letters
TRANSCRIPTION DNA to RNA Occurs in
nucleus Three process
Initiation Elongation Termination
RNA polymerase
DNA of gene
PromoterDNA Terminator
DNAInitiation
Elongation
TerminationGrowingRNA
RNApolymerase
Completed RNA
INITIATION RNA polymerase
binds to DNA Binds at
promoter region TATA box
RNA polymerase unwinds DNA
Transcription unit Part of gene that
is transcribed
INITIATION Transcription factors
bind to specific regions of promoter
Provide a substrate for RNA polymerase to bind beginning transcription
Forms transcription initiation complex
ELONGATION RNA molecule is built
RNA polymerase Primer not needed 5’ to 3’ 3’ to 5’ DNA is template
strand Coding strand
DNA strand that is not copied
Produces mRNA Messenger RNA
DNA double helix reforms
TERMINATION RNA polymerase recognizes a
termination sequence – AAAAAAA Nuclear proteins bind to string of
UUUUUU on RNA mRNA molecule releases from template
strand
POST-TRANSCRIPTIONAL MODIFICATIONS
Pre-mRNA undergoes modifications before it leaves the nucleus
Poly(A) tail Poly-A polymerase Protects from RNA
digesting enzymes in cytosol
5’ cap 7 G’s Initial attachment site for
mRNA’s to ribosomes Removal of introns
SPLICING THE PRE-MRNA DNA comprised of
Exons – sequence of DNA or RNA that codes for a gene
Introns – non-coding sequence of DNA or RNA
Spliceosome Enzyme that
removes introns from mRNA
SPLICING PROCESS Spliceosome contains a handful of small
ribonucleoproteins snRNP’s (snurps)
snRNP’s bind to specific regions on introns
ALTERNATIVE SPLICING Increases number and variety of
proteins encoded by a single gene ~25,000 genes produce ~100,000
proteins
TRANSLATION mRNA to protein Ribosomes read
codons tRNA assists
ribosome to assemble amino acids into polypeptide chain
Takes place in cytoplasm
TRNA Contains
triplet anticodon amino acid
attachment site Are there 61 tRNA’s
to read 61 codons?
TRNA: WOBBLE HYPOTHESIS First two nucleotides of codon for a specific AA is always
precise Flexibility with third nucleotide Aminoacylation – process of adding an AA to a tRNA
Forming aminoacyl-tRNA molecule Catalyzed by 20 different aminoacyl-tRNA synthetase
enzymes
RIBOSOMES Translate mRNA chains into amino acids Made up of two different sized parts
Ribosomal subunits (rRNA) Ribosomes bring together mRNA with
aminoacyl-tRNAs Three sites
A site - aminoacyl P site – peptidyl E site - exit
1 Codon recognition
Amino acid
Anticodon
AsiteP site
Polypeptide
2 Peptide bond formation
3 Translocation
Newpeptidebond
mRNAmovement
mRNA
Stopcodon
TRANSLATION PROCESS
Three stages Initiation Elongation Termination
INITIATION Ribosomal subunits associate with mRNA Met-tRNA (methionine)
Forms complex with ribosomal subunits Complex binds to 5’cap and scans for start codon (AUG) –
known as scanning Large ribosomal subunit binds to complete ribosome Met-tRNA is in P-site Reading
frame is established to correctly read codons
ELONGATION Amino acids
are added to grow a polypeptide chain
A, P, and E sites operate
4 Steps
TERMINATION A site arrives at a stop codon on mRNA
UAA, UAG, UGA Protein release factor binds to A site releasing
polypeptide chain Ribosomal subunits, tRNA release and detach from
mRNA
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Red object = ?What molecules are present in this photo?
POLYSOME
PROKARYOTIC RNA TRANSCRIPTION/TRANSLATION
Throughout cell Single type of RNA
polymerase transcribes all types of genes
No introns mRNA ready to be
translated into protein mRNA is translated by
ribosomes in the cytosol as it is being transcribed
REVIEW What is a gene? Where is it located? What is the main
function of a gene? Do we need our
genes “on” all the time?
How do we turn genes “on” or “off”?
REGULATING GENE EXPRESSION Proteins are not required by
all cells at all times Regulated Eukaryotes – 4 ways
Transcriptional (as mRNA is being synthesized)
Post-transcriptional (as mRNA is being processed)
Translational (as proteins are made)
Post-translational (after protein has been made)
Prokaryotes lacOperon trpOperon
TRANSCRIPTIONAL REGULATION Most common DNA wrapped around histones keep gene promoters
inactive Activator molecule is used (2 ways)
Signals a protein remodelling complex which loosen the histones exposing promoter
Signals an enzyme that adds an acetyl group to histones exposing promoter region
TRANSCRIPTIONAL REGULATION Methylation
Methyl groups are added to the cytosine bases in the promoter of a gene (transcription initiation complex)
Inhibits transcription – silencing Genes are placed “on hold” until they are needed E.g. hemoglobin
POST TRANSCRIPTIONAL REGULATION
Pre-mRNA processing Alternative splicing
Rate of mRNA degradation Masking proteins –
translation does not occur Embryonic development
Hormones - directly or indirectly affect rate
Casein – milk protein in mammary gland
When casein is needed, prolactin is produced extending lifespan of casein mRNA
TRANSLATIONAL REGULATION Occurs during protein synthesis by a
ribosome Changes in length of poly(A) tail
Enzymes add or delete adenines Increases or decreases time required to
translate mRNA into protein Environmental cues
POST-TRANSLATIONAL REGULATION Processing
Removes sections of protein to make it active Cell regulates this process (hormones)
Chemical modification Chemical groups are added or deleted Puts the protein “on hold”
Degradation Proteins tagged with ubiquitin are degraded Amino acids are recycled for protein synthesis
PROKARYOTIC REGULATION lacOperon
Regulates the production of lactose metabolizing proteins
PROKARYOTIC REGULATION trpOperon
Regulates the expression of tryptophan enzymes
CANCER Lack regulatory mechanisms Mutations in genetic code (mutagens)
Probability increases over lifetime Radiation, smoking, chemicals
Mutations are passed on to daughter cells Can lead to a mass of undifferentiated cells
(tumor) Benign and malignant
Oncogenes Mutated genes that once served to stimulate cell
growth Cause undifferentiated cell division
GENETIC MUTATIONS Positive and negative
Natural selection – evolution Cancer –death
Small-Scale – single base pair Point mutations
Substitution, insertion/deletion, inversion Large-Scale – multiple base pairs
SMALL-SCALE MUTATIONS Four groups
Missense, nonsense, silent, frameshift Lactose, sickle cell anemia
SNPs – single nucleotide polymorphisms Caused by point mutations
MISSENSE MUTATION Change of a single base pair or group of base
pairs Results in the code for a different amino acid Protein will have different sequence and structure
and may be non-functional or function differently
NONSENSE MUTATION Change in single base pair or group of
base pairs Results in premature stop codon Protein will not be able to function
SILENT MUTATION Change in one or more base pairs Does not affect functioning of a gene Mutated DNA sequence codes for same
amino acid Protein is not altered
FRAMESHIFT MUTATION One or more nucleotides are inserted/deleted
from a DNA sequence Reading frame of codons shifts resulting in
multiple missense and/or nonsense effects Any deletion or insertion of base pairs in
multiples of 3 does not cause frameshift
LARGE-SCALE MUTATIONS
Multiple nucleotides, entire genes, whole regions of chromosomes
LARGE-SCALE MUTATIONS Amplification – gene
duplication Entire genes are
copied to multiple regions of chromosomes
LARGE-SCALE MUTATIONS Large-scale deletions
Entire coding regions of DNA are removed Muscular Dystrophy
LARGE-SCALE MUTATIONS Chromosomal translocation
Entire genes or groups of genes are moved from one chromosome to another
Enhance, disrupt expression of gene
LARGE-SCALE MUTATIONS Inversion
Portion of a DNA molecule reverses its direction in the genome
No direct result but reversal could occur in the middle of a coding sequence compromising the gene
LARGE-SCALE MUTATIONS Trinucleotide repeat expansion
Increases number of repeats in genetic code CAG CAG CAG CAG CAG CAG CAG CAG
Huntingtons disease
CAUSES OF GENETIC MUTATIONS Spontaneous mutations
Inaccurate DNA replication Induced mutations
Caused by environmental agent – mutagen Directly alter DNA – entering cell nucleus Chemicals, radiation
CHEMICAL MUTAGENS Modify individual nucleotides
Nucleotides resemble other base pairs Confuses replication machinery – inaccurate copying
Nitrous acid Mimicking DNA nucleotides
Ethidium bromide – insert itself into DNA
RADIATION - LOW ENERGY UV B rays Non-homologous end joining
Bonds form between adjacent nucleotides along DNA strand
Form kinks in backbone Skin cancer
RADIATION – HIGH ENERGY Ionizing radiation – x-ray, gamma rays Strip molecules of electrons Break bonds within DNA
Delete portions of chromosomes Development of tumors
MUTATION IN PROKARYOTES DNA is mostly coding sequences Mutation is harmful – superbugs
GENOMES AND GENE ORGANIZATION Human Body
22 autosomal chromosomes 1 pair of each sex chromosome (XX, YY)
GENOMES AND GENE ORGANIZATION
Components VNTR’s–variable number tandem repeats
(microsatellites) Sequences of long repeating base pairs TAGTAGTAGTAGTAG
LINEs – long interspersed nuclear elements SINEs – short interspersed nuclear elements Transposons – small sequences of DNA that move
about the genome and insert themselves into different chromosomes
Pseudogene – code is similar to gene but is unable to code for protein
VIRUSES Not alive but can replicate themselves Contain
DNA or RNA Capsid – protein coat Envelope – cell membrane
VIRUS 4000 species of virus have been
classified
REPLICATION DNA
Transcription and translation RNA (retrovirus)
Uses reverse transcriptase – enzyme Uses cells parts to make a single strand of
DNA and then makes a complementary strand from that copy
Integrase – incorporates into our genetic code
VIRUS AS VECTORS Transduction
Using a virus vector to insert DNA into a cell or bacterium