from gene to protein lecture notes biol 100 – k.marr 1.topics for the next few lectures –...
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From Gene to Protein Lecture NotesBiol 100 – K.Marr
1. Topics for the next few lectures
– Transcription: From DNA to RNA
– Translation: From RNA to Protein
– Understanding Cystic Fibrosis
– Chapter 10 in Essential Biology by Campbell et al
2. Lab 7. Modeling DNA Structure, DNA Replication and Protein Synthesis Read the introduction carefully
– Part 1 (through page 9)—modeling DNA Structure and Replication
– Part 2—modeling transcription and translation
Cytoplasm
Nucleus
DNA
Transcription
Translation
Protein
mRNA
• Transcription: DNA copied into mRNA molecule
• Translation: ribosomes translate mRNA into protein—a chain of amino acids
• Proteins control phenotype. How?
The Flow of Genetic Information: DNA to RNA to Protein Phenotype
A few of the many roles played by proteins:
1. Enzymes: catalysts for nearly all chemical reactions in cells; Determine what cells can make and digest
2. Structural components: muscles (actin and myosin), connective tissue (collagen, elastin)
3. Receptors on cell surface for growth factors, hormones, etc.
4. Hormones: e.g. insulin, growth hormone, prolactin
5. Transport: e.g. hemoglobin, spindle fibers
6. Immune system: antibodies
The one gene–one protein hypothesis:The function of a gene is to dictate the production of a specific protein. Why are proteins so important?
CF phenotype
• Genes determine which proteins a cell can make
• Proteins control phenotype
• e.g. CFTR Gene codes for CFTR protein
CFTR Protein: The cystic fibrosis transmembrane regulator proteinCarbohydrate
Chloride ions
Water
CFTR Protein
Cellmembrane
Cytoplasm of cell lining duct or lungs
Water
Inside of duct or
Air sac in lungs
CFTR Protein
• Pumps chloride ions (salt) into cells lining ducts or the lungs
• What are the consequences when CFTR doesn’t work?
• How does a gene control the production of a protein?
The order of Bases in a gene determines the order of amino acids in the protein it codes for
Is the order of amino acids in a protein important?
• View animation of transcription
Questions to answer:
1. What do we start with and end with?
2. Where does transcription occur? When?
3. What is needed for transcription to occur?
4. What is the sequence of events?
Transcription: copying DNA into RNA
RNApolymerase
RNA nucleotides
Newly madeRNA Direction of
transcriptionTemplatestrand of DNA
Transcription of a gene by RNA polymerase
(a) Parent DNA
Strandseparation
(b) Transcription begins
RNApolymerase
Complementarybase pairing
Transcription: copying DNA into RNA ( 1 of 2)
(c) Transcription continues (d) Products of transcription
Non-codingstrand
Codingstrand
Nucleotidejoining
New RNA strand(actually severalhundred basepairs long)
Parent DNAtotallyconserved
Transcription: copying DNA into RNA ( 2 of 2)
Fig. 7.07
(a) Gene
(b) Primary transcriptTranscription
RNA splicing:
Differential splicing can result in different mRNA molecules and, therefore, different proteins
RNA Processing
Translation
(c) Spliced RNA
(d) Mature RNA
(d) protein
Exon 1 Exon 2
Intron 1
Exon 3
Intron 2
Exon 4
Intron 3
Exon 5
Intron 4
Exon 6
Intron 5
Transcription in Eukaryotic Cells: Differential RNA splicing can result in one gene producing more than one protein
Processing of Eukaryotic RNA
RNA Processing includes• Adding a cap and tail
• Removing introns
• Splicing exons together
– Differential splicing produces different mRNA molecules
Gene (DNA)
RNAtranscriptwith capand tail
mRNA
Exon Intron ExonIntron Exon
Cap
Introns removed Tail
Exons spliced together
Coding sequence
Nucleus
Cytoplasm
Transcription + theAddition of cap and tail
• View animation of translation
Questions to answer1. What do we start with and end with?
2. Where does translation occur?
3. What is needed for translation to occur?
4. What is the sequence of events?
5. What are the roles of mRNA, ribosomes, start codon, tRNA, anticodons, stop codon?
Translation: Ribosomes reading mRNA to produce a polypeptide
Transfer RNA: tRNA
tRNA1. Acts as a
molecular interpreter
2. Carries amino acids
3. Matches amino acids with codons in mRNA using anticodons
Amino acid attachment site
Hydrogen bond
RNA polynucleotide chain
AnticodonAnticodon
Amino acid
Codon on mRNAmRNA
A portion of an mRNA molecule attached to a tRNA
Each Codon codes
Specifies a specific
tRNA—amino acid
complex
Proteinunderconstruction
Largesubunit
Smallsubunit
A ribosome translating mRNA into protein
mRNA
Ribosomes• Organelle that makes
protein
• Reads mRNA 5’ 3’
• Made of rRNA and protein
• Consist of 2 subunits
Transcription & Translation of the CRTR Gene in Healthy People
Part of a normal CFTR gene:
5’...ATCATCTTTGGTGTT...3’ non-coding strand
3’...TAGTAGAAACCACAA...5’ coding strand
1. Transcribe this portion of the gene.
The whole gene codes for 1480 amino acids in CFTR protein!
What is the order of bases in the resulting mRNA molecule?
2. Translate this portion of the gene.
– What is the order of amino acids in the resulting protein?
Table of Codons
found on
mRNA
• Each codon specifies a specific amino acid
• The same genetic code is used by nearly all organisms!!
Part of a normal CFTR gene:
5’...ATCATCTTTGGTGTT...3’ non-coding strand
3’...TAGTAGAAACCACAA...5’ coding strand
5’...AUCAUCUUUGGUGUU...3’
Transcription
.....Ile-Ile-Phe-Gly-Val…
(only 5 of the 1480 amino acids in protein!!)
Translation
Transcription & Translation of the CRTR Gene in Healthy People
Part of CFTR gene associated with Cystic Fibrosis:
5’...ATCATTGGTGTT...3’ non-coding strand
3’...TAGTAACCACAA...5’ coding strand
1. Transcribe this portion of the gene.
What is the order of bases in the resulting mRNA molecule?
2. Translate this portion of the gene.
– What is the order of amino acids in the resulting protein?
3. What is different about the gene and the protein in people with cystic fibrosis?
Transcription & Translation of the CRTR Gene in People with CF
Part of CFTR gene associated with Cystic Fibrosis:
5’...ATCATTGGTGTT...3’ non-coding strand
3’...TAGTAACCACAA...5’ coding strandTranscription
Translation
Transcription & Translation of the CRTR Gene in People with CF
5’...AUCAUUGGUGUU...3’
.....Ile-Ile-Gly-Val…….. Phenylalanine (Phe) is missing
Explaining the symptoms of CF
• Why does CF only affect certain parts of the body?
• What do the characteristics of CF have in common?
1. Mucus build-up in the lungs
• Lung infections (e.g. pneumonia)
2. Male sterility (blocked vas deferens)
3. Salty sweat
4. Trouble digesting food (blocked pancreatic duct)
Explaining the symptoms of CF
Chloride ions in cell
Chloride ions outside of cell CFTR Protein: Pumps
Chloride ions into cell
• In CF, the faulty CFTR protein never makes it to cell membrane
1. What builds up outside of cells? Why?
2. Why salty sweat?
3. Why does mucus collect in lungs?
4. Why respiratory infections?
5. Why problems with digestion?
6. Why male sterility?
Understanding Cystic Fibrosis at the Cellular Level
How does CFTR protein get from where it’s produced to its home in the cell membrane?
1. Where is the CFTR protein produced?
2. CFTR is a glycoprotein—where does it go for modification?
How does it get there?
3. How does the modified CFTR protein get to the plasma membrane?
4. The defective CFTR protein is recognized at the ER as defective
Where is the defective CFTR protein sent?
CF symptoms may be mild or severe
Several hundred different mutations are associated with CF
CFTR Gene
What’s a Mutation?• Any change in the nucleotide sequence of DNA
• Types of Mutations
– Substitution, insertion or deletion
– Occur during DNA replication
• Mutations may Result from:
– Errors in DNA replication
– Mutagens
• physical or chemical agents that may cause errors during DNA replication
chemicals in cigarette smoke
Radiation (e.g. U.V. light, X-rays)
• Why does isoleucine (Ile) at amino acid position 507 remain unchanged?
F508 deletion: the most common cause of cystic fibrosis
Mutations responsible for Sickle Cell Anemia
Normal hemoglobin Sickle-cell hemoglobin
Glu Val
Normal hemoglobin DNA Mutant hemoglobin DNA
mRNA mRNA
• Only one amino acid in 146 is incorrect in sickle-cell hemoglobin!
Types of Mutations: Base Substitutions, Insertions or deletions
• Base substitutions
– May result in changes in the amino acid sequence in a protein, or
– May be silent (have no effect)
mRNA
Protein Met Lys Phe Gly Ala
(a) Base substitution
Met Lys Phe Ser Ala
Types of Mutations: Base Insertions and deletions
• Can have disastrous effects
– Change the reading frame of the genetic message
Met Lys Leu Ala His
(b) Nucleotide deletion
mRNA
Protein Met Lys Phe Gly Ala
• Although mutations are often harmful
– They are the source of the rich diversity of genes in the living world
– They contribute to the process of evolution by natural selection
DNA and RNA: Polymers of Nucleotides
SUMMARY OF KEY CONCEPTS
DNA
Polynucleotide
Nitrogenousbase
Phosphategroup
Nucleotide
Sugar
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