Unit 6 - Protein Synthesis Flashcards
Describe the structure of DNA
Purines: double ringed adenine and thymine
Pyrimidines: single ringed guanine and cytosine
5’ to 3’ order of nucleotide: phosphate group – 5-carbon sugar – nitrogenous base
The two strands of DNA are antiparallel.
10 base pairs for every DNA curl
Describe the wobble effect. How does it help?
Pairing of the tRNA anticodon with the mRNA codon: once the first two positions are paired, the third base of the anticodon can typically pair with not only a complementary base: it “wobbles”. For example, the double-ringed G can pair with either a single-ringed U or C.
This allows mRNA to be translated with fewer than the 64 tRNAs that would be otherwise be required. Some wobble positions can pair with any of the four bases.
Compare DNA and RNA
Compare mRNA, tRNA, and rRNA
Messenger RNA:
- Intermediary between DNA and ribosome
- Translated into protein by ribosome
- RNA version of gene coded by DNA
Transfer RNA:
- Functions as the delivery system of amino acids to ribosomes
- At least one type of tRNA for each amino acid
- Shaped like a 3-leaf clover
Ribosomal RNA:
- Binds with proteins to form the ribosomes
Before RNA leaves the nucleus, it is processed. What is an immature mRNA called? Explain this process
Immature mRNA: hnRNA (heterogenous nuclear)
A) 5’ Cap: the 5’ end is capped with a modified guanine nucleotide which serves as
- An attach here signal from the small ribosomal subunit
- Protection from enzymes
B) Poly-A Tail: 150 to 200 adenine nucleotides are added to the 3’ end which helps prevent degradation of the mRNA
C) Non-coding sequences (introns/intervening sequences) are removed. Exons/expressed sequences are kept
Describe the possible function of introns
- Plays a regulatory role in gene activity
- Allows different types of cells in the same organism to different proteins from a common gene
- Plays a role in the evolution of protein diversity
Where is the lac operon found?
In the E. coli within your intestines. It converts lactose that you drink into energy.
Descibe the layout of the lac operon
Left to right
Regulatory Gene: constantly transcribed and translated into an amino acid sequence that results in LacI repressor protein
Promotor: a DNA sequences that acts as a binding site for RNA polymerase
Operator: a binding site for LacI
Structural gene 1-3: these code for enzymes that break down lactose for energy
Explain how the lac operon is negatively controlled
Lactose absent:
- LacI binds to operator and blocks RNA polymerase from promoter
- Genes cannot be transcribed, no waste enzymes produced
Lactose present:
- Some lactose binds to LacI at control site, changing its shape and deactivating it
- RNA polymerase accesses promoter and produces enzymes
Explain how the lac operon is positively controlled
E. coli do not need lactose enzymes during high glucose concentrations because they utilize glucose instead.
Low glucose:
- cAMP increases, CAP binds and increases structual gene transcription
High glucose:
- cAMP decreases, CAP cannot bind and decreases structual gene transcription
Where is the trp operon found?
Normally in E. coli in the intestine, but for E. coli that exists elsewhere it must manufacture its own tryptophan
Explain how the trp operon is positively controlled
High tryptophan:
- Tryptophanbind to trp repressor protein at control site, altering its shape and allowing it to bind to operator, stopping tryptophan production
Low tryptophan:
- Trp repressor loses its shape without tryptophan binding, allowing RNA polymerase to bind to promoter and transcribe genes
What is meant by saying tryptophan is a corerepressor?
Corerepressor: a molecule that binds to a repressor to activate it.
Compare laq vs. trp operon
Lac operon is repressed during low lactose levels
Trp operon is repressed during high tryptophan levels
Point mutations: (3)
Point Mutations: changes in one or two nucleotides in a single gene.
Includes substitution, insertion, and deletion
What is the protein that breaks down mRNA when it is no longer needed?
Ribonuclease
Substitution:
Substitution: a replacement of one nucleotide and its partner with another pair of nucleotides. Sometimes has no effect due to redundancy and the wobble effect
Describe the two potential results from substitution
Missense mutation: the altered codon still codes for an amino acid and still makes sense but not always the intended sense
Nonsense mutation: the altered codon codes for premature termination (stop codon) creating a non-functional protein
Insertions and deletions:
Insertions and Deletions: addition or loss of one or more nucleotide
Duplication:
Duplication: a gene sequence in excess of its normal amount in a chromosome (ex., deletion from one chromosome is inserted in its homologue)
Inversion:
Inversion: a chromosome segment that separated from the chromosome and was reinserted at the same place but inverse
Translocation:
Translocation: the transfer of part of 1 chromosome (nonhomologous)
What is it called when a ribosome walks down the mRNA?
Translocation
Describe how chromosomes are packaged in the nucleus
- Every 200 nucleotides, the DNA is coiled around a core group of stabilizing proteins called histones (+ive) which are attracted to -ive DNA.
- This complex of histones enveloped by coiled DNA is called a nucleosome
- Next, the series of nucleosomes coil into chromatin fibres
- These fibres fold into the final chromatin structure via supercoiling
- One DNA helix forms each chromosome
How do you determine how many cuts with be made by a restriction enzyme in a set of DNA?
To tell how many cuts: 75000 (or simply the number of nucleotides in the DNA) / 4^(length of nucleotide to be cut).
Explain transcriptional control
- Regulate which genes are transcribed
- Control the rate at which transcription occurs.
Explain posttranscriptional control
- Converting hnRNA to mRNA
Explain translational control
- Controls how often and how rapidly mRNA transcripts will be translated into proteins.
- Affects the length of time it takes for mRNA to be activated
- Affects the speed at which enzymes destroy mRNA.
Explain posttranslational control
- Before many proteins become functional, they must pass through the cell membrane. Several control mechanisms affect this rate.
How many hydrogen bonds do the nitrogenous bases have?
Adenine and thymine have 2, guanine and cytosine have 3. So, adenine and thymine are easier to separate.
Define biotechnology
Biotechnology: the use of biological substances to develop an agricultural, medicinal, environmental, etc. product or process (synthetic hormones, insulin, food)
What is recombinant DNA?
Provide natural examples.
Recombinant DNA refers to new genetic material constructed from DNA of different organisms
Natural examples (evolution):
- Combinations of genes between different organisms (fertilization/crossing over)
- Bacteria: during transformation, they picked up free DNA
- Virus: sometimes transfers genes into eukaryotic cell host
State 3 tools used to create recombinant DNA in a lab. What are methylases?
Plasmids: ring of DNA found in bacteria that carries accessory genes which replicate independently of the useful DNA
DNA Ligase: an enzyme used to “glue” fragments by condensation that have been cut by the same restriction enzyme
Restriction Enzyme: enzymes that protect bacteria from bacteriophage DNA injections, make palindromic cuts.
- Methylases: enzymes that modify recognition sites by placing a methyl group on a base. This prevents the cutting of that portion of the DNA; prevents digestion of bacteria’s own DNA
Explain how plasmids, ligase, and restriction enzymes are used together
- Combining these three devices allows for the duplication of human DNA
- Plasmid and the human DNA sample are treated with the same restriction enzyme.
- The enzyme cuts out the specific sequence of the human DNA which is then inserted into the plasmid, replacing the plasmid DNA that was snipped as well.
- Ligase seals the ends. The bacteria are allowed to replicate.
What is gel electrophoresis? Describe the steps it requires
Gel electrophoresis is used to separate charged molecules on the bases of size through a gel meshwork. DNA is negatively charged due to its phosphate groups.
Steps
- DNA is cleaved by restriction enzymes into fragments of different lengths
- A gel is used which is comprised of a buffer and agarose
- A current causes DNA to move from the negative to positive terminal. Smaller fragments travel faster
What is RFLP
Restriction Fragment Length Polymorphism: any difference in DNA sequence that can be detected between individuals
Explain southern blotting. What is the x-ray used called?
A procedure that allows DNA in a gel to be transferred to a nylon membrane. A membrane is placed on the gel and the DNA is blotted onto it (positive electrode behind membrane). The DNA is then stained. The resulting picture generated via x-ray is called an autoradiogram.
Explain the process of PCR
Polymerase chain reactionis used to make many copies of a specific DNA region in vitro.
- PCR relies on a DNA polymerase, Taq polymerase
- The experimenter determines the region of DNA that will be amplified by the primers she or he chooses.
- The DNA primers must be complementary to the target area.
- When the primers are bound to the template, they are extended by Taq polymerase.
- The new DNA that’s made in one round can serve as a template in the next round of DNA synthesis.
Describe chain termination. What is it also known as?
A.k.a. DNA Sequencing or Sanger Dideoxy Method
- Treat double stranded DNA to become single stranded
- Add primer to end of DNA
- Add 4 identical copies of DNA to 4 tubes with each containing DNA polymerase, free nucleotides, and 1 dideoxy nucleotide (a nucleotide – A’, G’, C’, T’ – missing an OH on the 3’ carbon deoxyribose sugar, stopping DNA synthesis)
- This creates many different fragments of DNA which are organized via gel electrophoresis and are read off from bottom to top to get the complementary strand which is again read off to get the original strand.
