DNA and RNA Functions and Structure

• Nucleic acids include DNA and RNA.

Functions of DNA

• DNA (deoxyribonucleic acid) carries the genetic instructions for development, functioning, and reproduction in living organisms and some viruses.

Functions of RNA

• RNA (ribonucleic acid) can carry genetic information in viruses.
• Three types of RNA convert DNA code into polypeptides, or proteins.
• RNA can act as a catalyst, like enzymes.
• RNA has complex regulatory roles in cells, like gene expression and modifying other RNA.

Structure of Nucleic Acids

• A nucleotide monomer consists of a sugar, a nitrogenous base, and a phosphate group.
• Nitrogen-containing bases project from the backbone and are linked to a carbon in the sugar via a glycosidic bond.
• DNA is a large polymer, and human chromosome 1 has 249 million base pairs.

The Sugars in DNA and RNA

• Both DNA and RNA contain 5-carbon sugars.
• DNA contains deoxyribose.
• RNA contains ribose.
• A phosphate group at the 5' carbon bonds to the 3' carbon of the next sugar.
• Nitrogenous bases bond to the 1' carbon.

DNA Strands

• DNA strands are antiparallel and run in opposite directions.
• One strand runs 5' to 3', while the other runs 3' to 5'.
• There are 5 sugar bonds to each phosphate group.
• Each sugar also has a 3' hydroxyl group.
• DNA is most stable as a double-stranded helix.

Packaging DNA

• In eukaryotes, DNA is wrapped around proteins called histones, in a coiled form called chromatin.
• Chromatin is further compressed by supercoiling, forming highly compacted structures called chromosomes.
• Most prokaryotes lack histones but have supercoiled DNA held together by special proteins.

DNA Replication

• DNA replication involves separating DNA, building a complementary strand, and quality control with DNA repair.

Separating DNA

• DNA helicase enzymes unzip DNA.
• Helicase breaks hydrogen bonds between complementary base pairs.
• The junction that is created is called a replication fork.

Problem and Solution in DNA Replication

• To prevent hydrogen bonding between base pairs, single-stranded binding proteins (SSBs) bind to exposed DNA single strands.
• Topoisomerase regulates the unwinding of the DNA.

Building Complementary Strands

• DNA polymerase creates a new strand by pairing complementary bases to the separated template strands.
• DNA polymerase travels in the 3' to 5' direction on the template strand and builds the complementary strand in the 5' to 3' direction.
• DNA polymerase cannot initiate a new strand, so primase adds a short segment of nucleotides called a primer (short RNA sequence) to the 3' end of the template strand.
• DNA polymerase removes the primers and replaces them with DNA.
• Primase belongs to the family of enzymes called RNA polymerase.

DNA Polymerase

• DNA polymerase is a family of enzymes involved in DNA replication, containing 7 different subgroups.
• DNA polymerases I, II, and III are specific enzymes from E. coli where DNA replication was first studied.

Leading Strand

• The leading strand follows the 3' to 5' template strand and moves towards the replication fork, being replicated continuously.

Lagging Strand

• The lagging strand moves from the replication fork to the 5' end of the template strand, so replication is discontinuous.
• The resulting fragments are called Okazaki fragments.
• DNA ligase catalyzes the reaction to join the fragments together.

Semi-Conservative Replication

• Replication is called semi-conservative.
• This is because, one half of the original strand is always saved or conserved.

Genes

• A gene is a sequence of DNA that codes for a protein.
• Protein coding genes account for less than 2% of the nucleotides in the genome.
• Each 3-nucleotide base pair codes for an amino acid and is referred to as a codon.
• In addition to coding for a protein, other portions provide additional information, including sequences that control where and how much protein to make.

Universal Genetic Code

• Almost all protein-coding regions begin with the sequence AUG.
• AUG encodes for the amino acid methionine (Met).
• Three "stop" codons mark the end of the protein coding region.
• Multiple codons can code for the same amino acid.

Central Dogma

• Transcription copies information in DNA to messenger RNA (mRNA).
• Translation involves ribosomes reading the mRNA and assembling amino acids in sequence to form a protein or polypeptide.

Transcription

• Messenger RNA (mRNA) goes through transcription to translation.
• Transfer RNA (tRNA) carries the amino acids corresponding to the codon.
• The anticodon lines up on that codon and release an amino acid.

Transcription

• During transcription, information stored in DNA is copied to RNA.
• Only one DNA strand serves as a template for transcription at any given time.
• The template strand of DNA is also referred to as the non-coding strand.
• The non-template strand is referred to as the coding strand because its sequence will be the same as the new messenger RNA (mRNA molecule).

Three Stages of Transcription

• Initiation: RNA polymerase binds to template DNA.
• Elongation: RNA polymerase reads DNA and adds nucleotides to the 3' end of mRNA.
• Termination: RNA synthesis ceases and RNA polymerase and mRNA are released.
• Transcription is less complex in prokaryotes than eukaryotes.

Initiation in Eukaryotic Transcription

• RNA polymerase attaches upstream of a gene at a specialized sequence called a promoter.
• A promoter region usually consists of a string of T and A's, and then the DNA helix begins to unwind.

Elongation in Eukaryotic Transcription

• The double helix unwinds, and RNA polymerase reads the template strand.
• Then, added nucleotides get moved to the mRNA at the 3' end.
• The DNA that has been transcribed reforms the double helix.

Termination in Eukaryotic Transcription

• RNA polymerase reaches a terminator sequence in the gene.
• mRNA and RNA polymerase are cleaved.
• The mRNA will travel to a ribosome, and RNA polymerase can read another gene.

Modification After Transcription

• A 5' cap helps in the initiation of translation.
• A poly-A tail protects the mRNA from enzymes in the cytoplasm.
• A gene has exons (segments that code for protein) and introns (non-coding regions).
• The introns are removed, and the exons are joined together by spliceosomes (RNA and protein).

Alternate Splicing

• During alternate splicing, exons can be joined in different combinations.
• This allows for the production of different mRNAs.
• The mRNAs can then be translated to produce related proteins.

Protein Synthesis

• Translation is the process during which a protein is synthesized from the information in the mRNA.

3 Stages of Translation

• Initiation
• Elongation
• Termination

Ribosomes

• Ribosomes are organelles that consist of proteins and RNA.
• Translation occurs in this organelle.

Transfer RNA (tRNA)

• tRNA is a small, single-stranded RNA molecule.
• One arm contains an anticodon sequence of 3 bases that are complementary base pairs to mRNA, with the sequence being read.

Initiation in Translation

• Begins when the small subunit of a ribosome attaches to the cap (5' end) of mRNA and then moves along the mRNA to the initiation site.
• tRNA binds through an anticodon to the start codon of mRNA, typically AUG for methionine.
• The large ribosomal unit binds and forms the E (exit), P (peptidyl), and A (aminoacyl) sites.

Elongation in Translation

• The first tRNA occupies the P site.
• The 2nd tRNA enters the A site.
• Amino acids from the P site bind to the amino acid at the A site.
• The ribosome moves along.
• The 1st tRNA exits, and the tRNA moves to the P site.
• Now the new tRNA enters the A site, and the process continues as above, building the polypeptide strand.
• At the E site, tRNA that has no amino acid is released.

Termination in Translation

• Termination happens when a stop codon is encountered.
• Then the release factor enters the A site.
• The ribosome dissociates, and the newly formed polypeptide is released.

Polypeptide to Proteins

• To convert a polypeptide to a protein, it needs to be folded (3D shape).
• For quaternary proteins, multiple polypeptides need to be assembled.

Gene Regulation

• Cells must have the ability to turn genes on and off.
• Also, adjust the volume of each gene and produce the required amount of proteins.

Prokaryote vs Eukaryote

• Prokaryotes use a feedback model called an operon system.
• An operon is a cluster of coregulated genes that share a promoter and operator.
• Gene control is more complex in eukaryotes than prokaryotes.

Operon System

• Promoter - Site where DNA transcription occurs.
• Operator - Sequence of bases that control transcription; where repressor binds, stopping the transcription of the gene.
• Structural Genes - Genes coding for the protein(s) get transcribed as a unit.

Lac Operon

• The lac operon produces 3 proteins.

  • β-galactosidase splits bonds in lactose.
  • Galactoside permease is a transport protein that embeds in the cell membrane and pumps lactose into the cell.
  • Transacetylase

• The lac repressor is always present, so genes are normally off.

• Repressor is bound to operator region, thus RNA polymerase cannot transcribe DNA.

• Lactose (inducer) binds to repressor protein and is released from DNA.

• RNA polymerase reads DNA, and mRNA gets translated into proteins needed for lactose metabolism.

• When lactose levels increase, the repressor binds to the operator region again, stopping production of proteins.

TRP Operon

• Genes are normally on and produce the amino acid tryptophan.
• If tryptophan is present, it binds to the repressor and prevents more tryptophan from being made.

Eukaryotic Gene Regulation

• Transcriptional Regulation (Copying mRNA).
  • Regulates which genes are transcribed or the rate of transcription.
  • Access to promoters is either enhanced or decreased.
  • Example: Methylation of genes, transcription factors can't bind.
• Post-Transcriptional (After Copying mRNA).
  • Controls the availability of mRNA to ribosomes.
  • Example: Alternative splicing enables perhaps 75% of human genes to undergo this.
• Translational (Protein being Synthesized).
  • Controls how often and how mRNA is translated to a protein.
  • Example: Variation of the length of the poly(A) tail can regulate the efficiency of translation.
• Post-Translational (After Protein Synthesized).
  • Controls when proteins become functional, how long they are functional, and when they are degraded.
  • Example: Activation of P53, a tumor suppressor protein, occurs through phosphorylation.

Mutations

• Mutations are changes in the DNA.
• There are two classes of mutations: small scale and large scale.

Small Scale Mutations

• Point mutations occur with a single base change.
• A change of small group pairs may occur.
• Types of mutations:
  • Substitution: Replacement of one base by another
  • Insertion: Placement of one or more extra nucleotides in the sequence.
  • Deletion: Removal of one or more nucleotides in the sequence.
  • Inversion: Two adjacent bases trade places.
• Deletion and insertion can cause changes to reading frames of codons.
• Effects of mutations:
  • Silent: Does not change the amino acid coded, and thus has no effect.
  • Nonsense: Converts codon into a stop signal.
  • Missense: Results in the single substitution of one amino acid.

Frame Shift Mutation

• Insertion or deletion of one or more base pairs causes the reading frame to shift in one direction or the other.
• These result in multiple missense and/or nonsense effects.

Large Scale Mutations

• These mutations may involve:
  • Large chunks of DNA inserted, lost, or repeated.
  • Duplication or loss of genes.
  • Whole regions of chromosomes.

Causes of Mutations

• Spontaneous mutations: Result from errors during DNA copying, or damage to bases but are not caused by factors from the external environment.
• Induced mutations: Damage caused by exposure to mutagenic agents, such as UV radiation, x-rays, or certain chemicals.

Example of Mutation

• Sickle Cell Anemia has a mutation that alters a single nucleotide, so the sixth amino acid becomes valine instead of glutamic acid.
• This changes the shape of protein hemoglobin, causing them to form rigid rods that stick together and can intercept blood flow.

Importance of Mutations

• These provide raw material for genetic diversity and are essential for evolution.
• Mutations can be beneficial, harmful, or neutral.
• Mutation rates may be increased by the external environment.
• However, they are random, in that whether a particular mutation happens or not is unrelated to how useful that mutation would be.