Molecular Genetics Test Flashcards
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nucleotide structure
Each nucleotide consists of:
A phosphate group
A deoxyribose or ribose sugar
A nitrogenous base (adenine, thymine, cytosine, guanine)
bases, pyrimidines vs purines
Pyrimidines: Cytosine (C) and Thymine (T) (one-ring structure)
Purines: Adenine (A) and Guanine (G) (two-ring structure)
Complementary base pairing and Chargaff’s rule
Complementary Base Pairing
A pairs with T (two hydrogen bonds)
C pairs with G (three hydrogen bonds)
Chargaff’s Rule
States that in any given DNA molecule, the amount of adenine equals thymine, and the amount of cytosine equals guanine (A=T, C=G).
all enzymes and their functions (terms: replication fork, SSBPs, helicase, topoisomerase (gyrase); primase, DNA polymerase III, DNA Polymerase I, ligase
SSBPs (Single-Strand Binding Proteins): SSBPs are proteins that bind to single-stranded DNA during replication, to help stabilize the newly unwound single DNA strands.
Replication Fork: The replication fork is the Y-shaped structure formed during DNA replication where the DNA is unwound and the two strands are separated to allow for the synthesis of new DNA strands.
DNA helicase - unwinds and separates double stranded DNA as it moves along the DNA. It forms the replication fork by breaking hydrogen bonds between nucleotide pairs in DNA.
DNA primase - a type of RNA polymerase that generates RNA primers. Primers are short RNA molecules that act as templates for the starting point of DNA replication.
Topoisomerase or DNA Gyrase - relieves tension in DNA strands to prevent the DNA from becoming tangled or supercoiled.
DNA polymerases - synthesize new DNA molecules by adding nucleotides to leading and lagging DNA strands.
DPI → Replaces RNA primers with DNA nucleotides and proofreads and corrects typos in the DNA
DPIII → adds DNA nucleotides in the 5’ to 3’ direction
DNA ligase - joins DNA fragments together by forming phosphodiester bonds between nucleotides.
Describe the Process of DNA Replication (Initiation)
Unwinding the DNA: The process begins at specific locations called origins of replication. The enzyme helicase unwinds the double helix, creating a replication fork. Single-stranded binding proteins (SSBPs) stabilize the unwound DNA strands.
Relieving Tension: As DNA unwinds, topoisomerase alleviates the tension ahead of the fork, preventing supercoiling.
Primer Synthesis: The enzyme primase synthesizes short RNA primers complementary to the template strands, providing starting points for DNA synthesis.
Describe the Process of DNA Replication (Elongation)
Nucleotide Addition: DNA polymerase III attaches to the RNA primer and begins adding nucleotides in the 5’ to 3’ direction. On the leading strand, this occurs continuously.
Lagging Strand Synthesis: On the lagging strand, DNA is synthesized in short segments called Okazaki fragments. Each fragment is initiated by a new RNA primer.
Primer Replacement: Once DNA synthesis is complete, DNA polymerase I removes the RNA primers and replaces them with DNA nucleotides.
Sealing Gaps: The enzyme ligase then seals the gaps between Okazaki fragments (using phosphodiester bonds), ensuring a continuous DNA strand.
Describe the Process of DNA Replication (Termination)
-Occurs upon completion of the 2 new strands of DNA are synthesized, 2 double strand DNA molecules are produced that will recoil
into a helix.
- replication machine is dismantled
Replication Models
1) Conservative: The original DNA molecule remains intact, and a completely new molecule is synthesized.
2) Semi-conservative: the original molecule is split in half, and the other side is filled-in
3) Dispersive: Each new molecule is comprised of bits and pieces of both new DNA and the original strand
Meselson-Stahl Experiment:
The Meselson-Stahl experiment demonstrated that DNA replication is semi-conservative.
Setup: E. coli bacteria were grown in a medium containing the heavy nitrogen isotope N-15, labeling their DNA.
Transfer: The bacteria were then moved to a medium with the lighter isotope N-14.
Sampling: After one and two rounds of replication, samples of DNA were taken.
Analysis: The DNA was separated by density using centrifugation.
Results:
- After one replication, the DNA had an intermediate density (one old strand and one new).
- After two replications, there were both light DNA (two N-14 strands) and intermediate DNA (one N-15 and one N-14 strand).
This proved that the Semi-conservative model was correct
Role of telomeres and telomerase
Telomeres are repeating sequences of nucleotides
- If chromosomes were not capped by telomeres, a small portion of a gene near the end of the chromosome could be lost every time DNA replication occurred
- Instead, only portions of telomeres are lost
Telomerase: An enzyme that extends telomeres, particularly active in germ cells and some stem cells, helping maintain chromosome integrity.
Germ line cells are unique because they must be able to continue replicating-
- Must maintain genetic integrity from parent to offspring
-Enzyme called telomerase adds more DNA to shortening telomeres, restoring their length
-Stem cells and some white blood cells also show presence of telomerase
RNA vs DNA
DNA:
Sugar: DNA contains deoxyribose, a sugar with one less oxygen atom than ribose.
Strands: DNA is typically double-stranded, forming a double helix.
Bases: The four nitrogenous bases in DNA are adenine (A), thymine (T), cytosine (C), and guanine (G). Adenine pairs with thymine, and cytosine pairs with guanine.
Shape: The double-stranded structure is stable and coiled into a double helix.
RNA:
Sugar: RNA contains ribose, a sugar with one more oxygen atom than deoxyribose.
Strands: RNA is usually single-stranded.
Bases: RNA uses adenine (A), uracil (U), cytosine (C), and guanine (G). In RNA, uracil replaces thymine as the complementary base to adenine.
Shape: RNA is typically single-stranded and can fold into various shapes, depending on its function.
- Function
DNA:
Genetic Material: DNA stores the genetic instructions used in the growth, development, functioning, and reproduction of all living organisms. It is the blueprint for all biological information.
Replication: DNA is replicated when cells divide, ensuring that each new cell receives an exact copy of the genetic information.
RNA:
Protein Synthesis: RNA plays a central role in protein synthesis.
Transcription (Initiation)
Promoter region: a sequence of nucleotides in DNA that indicates where the RNA polymerase complex should bind to initiate transcription
-Key element of the promoter in eukaryotes is the TATA box (a portion of DNA with high percentage of Adenine and Thymine bases)
-Prokaryotes have a TATATT sequence instead for this
-RNA polymerase binds to a promoter region on the DNA same purpose
Transcription (Elongation)
RNA polymerase complex works its way along DNA molecule
-Without needing a primer to be already in place
-Synthesizes mRNA strand that is complementary to template strand of DNA (T is replaced with U)
-RNA polymerases work in the 5ʼ → 3ʼ direction, using the 3ʼ→ 5ʼ DNA strand as a template strand.
As RNA polymerase moves along the DNA, it unwinds the DNA at the forward end of the enzyme
-RNA strand grows as nucleotides are added, one by one forming a temporary RNA-DNA double helix with the template strand
-As the RNA polymerase passes, the DNA double helix reforms
-Once an RNA polymerase molecule has started transcription and progressed past the beginning of a gene, another molecule of RNA polymerase may start producing another RNA molecule if there is room at the promote
Transcription (Termination)
The transcription is terminated when RNA polymerase recognizes a termination sequence.
Them RNA has been completely transcribed
●In eukaryotes, this is pre-mRNA and must be further processed before exiting the nucleus
Coding vs Template strands (which goes in which direction)
Coding (sense) is the same sequence as the created mRNA strand but with thymine instead of uracil 5’ to 3’ direction
Template (anti-sense) is the strand used as the template for the mRNA strand 3’ to 5’ direction
mRNA processing in eukaryotes (poly A tail, G cap, splicing of exons
Poly(A) tail: a chain of adenine nucleotides added to the 3’ end of pre-mRNA molecule to protect it from enzymes in the cytosol
-Enables mRNA to be translated efficiently and protects from attack by RNA-digesting enzymes in the cytosol
Splicing
The intron sequences are removed from pre-mRNA and exons are joined together to form mature mRNA
G Cap
-Involves covalent linkage of modified G nucleotides onto to 5’ end of pre-mRNA
-The cap is recognized by the protein synthesis machinery
-All eukaryotic mRNA undergo modifications on their ends
-These modifications convert precursors mRNA (pre-mRNA) to mature mRNA
Exons vs introns, alternate splicing
Exons are coding regions
Alternate Splicing
Exons may be joined in different combinations to produce different mRNAs from a single DNA gene sequence
-A mechanism called alternative splicing increases number and variety of proteins encoded by a single gene
-Allows more than one possible polypeptide to be made from a single gene
-Alternative splicing helps understand why humans with only 20 000 genes can produce approx. 100 000 proteins
Structure of tRNA (anticodon
-3’ end of tRNA binds to specific amino acids
-Anticodon on tRNA complements mRNA codon
Structure of ribosome - a, p and e sites
-2 subunits: small and large
-E site → exit site
-P site → polypeptide binding site
-A site → amino acid site
-Composed of proteins and rRNA
-3 tRNA binding sites
Translation - Initiation
-mRNA, tRNA and small ribosomal subunit bind with P site at start codon
-Only the first tRNA binds to P at start codon
-Large subunit binds using energy from GTP
Translation - Elongation
mRNA read 3 nucleotides at a time in codons
-tRNA brings corresponding amino acid into the A site of the ribosome
-Ribosome catalyzes dehydration synthesis reaction between amino acids in P site and A site
-Peptide bond is formed
-Growing polypeptide now attached to tRNA in A site
-Ribosome moves forward one codon
-Free tRNA in P site exits out the back of ribosome on the E site
-tRNA (with polypeptide) moves into P site
Translation - Termination
-Elongation continues until reaching a stop codon
-There is no amino acid for stop codon, just STOP
-Release factor binds and hydrolyzes the bond between the last tRNA and its amino acid
-New protein is free
frameshift vs. point mutations, missense, nonsense, silent
Frameshift Mutations
– shifts the reading frame of the codons so that a completely different amino acid sequence is produced from the point of the shift. Will likely not produce a functional protein
point mutations
– change in one nucleotide
○Substitution
○Substitutions will only affect a single codon
■THE FAT CAT ATE THE RAT
■THE FAT HAT ATE THE RAT
○Insertion (Frameshift)
■THE FAT CAT ATE THE RAT
■THE FAT CAT HATE THE RAT
○Deletion (Frameshift)
■THE FAT CAT ATE THE RAT
■THE FAT CAT AT THE RAT
Missense
A substitution that results in a different amino acid is
called a missense mutation
Nonsense
A nonsense mutation produces a premature STOP codon
Silent
If the mutation still produces the same amino acid it is
called a silent mutation. -Has no impact on the protein formed-This happens because the genetic code is degenerate
Structure of an operon and role of components (promotor, operator, repressor, genes)
Structure of an Operon:
Promoter: A DNA sequence where RNA polymerase binds to initiate transcription of the operon.
Operator: A DNA region that acts as a switch. It is located between the promoter and the structural genes, and it is where a repressor protein can bind to block transcription.
Repressor: A protein that can bind to the operator and prevent transcription. The repressor is often regulated by an inducer or co-repressor molecule.
Structural genes: These are the genes that code for proteins, often enzymes, that carry out a particular function (e.g., breaking down a sugar or synthesizing an amino acid).
