Nucleic Acids, DNA Replication, and Protein Synthesis

Posted on Feb 5, 2025 in Biology

Nucleic Acids

DNA Functions: Stores genetic information, transmits information to other molecules and between generations. 10 base pairs per turn.

Nucleotides: Sugar and phosphate make the backbone; bases give chemical identity. Bases: Adenine + Thymine (2 H-bonds), Guanine + Cytosine (3 H-bonds). Has 1-3 phosphate groups attached. A, G = purines. T, C = pyrimidines.

Chargaff’s Base Pairing Rule: A-T, C-G (helped discover the helical shape of DNA). Pair because of base distance, H-bond pattern, and stacking interactions above and below.

Nucleoside: JUST sugar and base.

Phosphodiester Bond: Connects C-O-P-O-C. Twists form uneven grooves: major and minor (antiparallel).

Base Stacking: Stabilizing van der Waals forces.

RNA: Has a ribose sugar instead of deoxyribose (1 more OH – less stable, more highly reactive). Uracil instead of Thymine. Shorter and single-stranded. 5′ DNA is monophosphate, 5′ RNA is triphosphate.

Levels of Structure:

  • Primary – 1 strand
  • Secondary – 2 strands
  • Tertiary – chromatin (wound up DNA)

If AT and CG are mismatched: bulging or spacing occurs. The hydrophobic effect causes the formation of the helix.

Biological Information

Central Dogma: Biological information flow (DNA to mRNA to polypeptide).

DNA Replication (duplication of DNA in a cell) + Protein Synthesis (synthesis of RNA/Proteins)

Replication: DNA makes exact copies to pass to cells or progeny. Separate into 2 strands, each undergoes synthesis of a complementary daughter strand. Errors can lead to mutations.

Protein Synthesis:

  1. Transcription: DNA to RNA
  2. Translation: RNA to amino acids (Prokaryotes – both in cytoplasm, Eukaryotes – transcription in the nucleus)

rRNA: Not translated, single strands of RNA assembled with proteins, integrated into ribosomal proteins. Eukaryotes: concentrated in the nucleus, Bacteria: everywhere

tRNA: Not translated, folded into a structure for translation. Carries individual amino acids.

Both of the above are considered stable because they are bonded or folded with protein.

mRNA: Translated into protein in protein synthesis. Unstable: RNases that degrade RNA are present everywhere (easy to do because it is single-stranded).

Gene: Transcription unit contains:

  1. Promoter
  2. Complete sequence of transcript (RNA)
  3. Transcription terminator sequence

Polymerases and enzymes move along DNA in the 3′ to 5′ direction (mRNA is built 5′ to 3′).

RNA is antiparallel to the template strand, the same as the coding strand.

Transcription

  1. Initiation: RNA polymerase and proteins are attached to the DNA, and the DNA is separated (+1 site is the first transcribed site).
  2. Elongation: Nucleotides are added to the 3′ end of RNA. Energy from broken DNA bonds is used for RNA bonds.
  3. Termination: RNA encounters a “stop sequence” and is released after RNA polymerase passes through the terminator.

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Eukaryotes have 3 kinds of RNA Polymerase, Bacteria have 1.

RBS is for translation but is still present on DNA. Same with start and stop codons.

Sigma Factor: Mediates promoter recognition in bacteria. Once transcription starts, it dissociates. Sigma factor + RNA polymerase = HOLOENZYME.

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TBP: TATA-binding protein helps RNA polymerase bind. The transcription start site is 25 bases from TATA.

General Transcription Factors: Assemble the promoter of a gene in eukaryotes. TBP + GTF = Basal Transcription Complex (minimum needed for transcription in eukaryotes).

Transcriptional Activator Protein: Binds to an enhancer to allow transcription to begin.

Enhancer: Specific DNA sequence necessary for transcription.

Hairpin Loop: Forms on bacterial mRNA at the termination point.

Transcription Details

RNA Polymerase: Moves along the 3′ to 5′ strand. New RNA is 5′ to 3′. Made up of protein subunits. Binds to minor and major grooves. Recognizes specific base pair sequences. Binds with H-bonds and non-covalent interactions. R groups interact with bases (different for AT and CG).

Bacterial Transcription: The promoter has a -10, -35 box. The sigma factor binds to boxes and helps RNA polymerase bind. The holoenzyme knows the orientation by moving in the -10 direction. The ability to bind tightly to the promoter is a level of control. Starts transcription at the +1 site. Stops after the termination sequence; a hairpin loop forms on mRNA, which causes RNA polymerase to dissociate.

Eukaryotic Transcription: 5′-TATAA-3′ (-25). TBP initiates transcription (helps RNA polymerase bind). The basal transcription complex binds to the promoter. Starts transcription at the transcription start site. Stops after the termination sequence, involves “termination factors” (proteins) and multiple repeats of bases.

The polymerization reaction is made irreversible by hydrolysis of the pyrophosphate group.

Different genes can be transcribed based on the orientation of the promoter.

Primary Transcript: RNA transcript off of DNA. mRNA when combined with a ribosome. In prokaryotes, the two are the same; ribosomes bind and begin protein synthesis immediately. In eukaryotes, processes are separated by the nucleus. This allows for RNA processing – modification of the primary transcript:

  1. 5′ cap added (RBS)
  2. Polyadenylation (addition of poly A tail -AAAAA)
  3. Removal of introns by spliceosome

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Alternative Splicing: Primary transcript is sliced in different ways to make different mRNA.

NTPs in Polymerization (Activated Monomers):

  • NTP – nucleotide triphosphates
  • dNTPs – DNA monomers
  • rNTPs – RNA monomers

They provide energy.

ATP Hydrolysis gives energy to drive polymerization and transport. It is spontaneous and energetically favorable. Release of phosphates from ATP.

DNA Packaging:

  • Bacteria – promoters unpackaged, always default + accessible to RNA polymerase
  • Eukaryotes – packaged tight, must be decondensed before transcription

ORF: Open reading frame, all bases translated in translation (start to stop codon)

Translation

Translation requires:

  • Ribosomes:
    • Large subunit – A (aminoacyl), P (peptidyl), E (exit) sites
    • Small subunit – assures mRNA splits into 3-letter reading frames
  • tRNA – Translation of codons to amino acids
  • Aminoacyl tRNA Synthetases: Connect amino acids to tRNA
  • Genetic Code: Correspondence between codons and amino acids
  1. Initiation: Initiation factors recruit small ribosomal units and tRNA and scan. Initiation factors are released, large ribosomes join, the next tRNA binds to the A site, and the original tRNA is in the P site. The reaction attaches the P-site codon to the A-site (peptide bond). The codon is recognized, and the first amino acid is created in the polypeptide chain. Requires protein initiation factors (eukaryotes – 5′ cap. Bacteria – Shine-Dalgarno sequence).
  2. Elongation: The ribosome moves down, and the original tRNA is ejected through the E-site. Successive amino acids are added. Elongation factors: bound to GTP, break their bonds to provide energy for elongation.
  3. Termination: Addition of amino acids stops, and the chain is released.

tRNA moves through the A-site to the P-site to the E-site. Protein is made H2N—COOH like RNA is 3′ to 5′.

Proteins

Allosteric:

  1. Confirmation change
  2. Change enzyme shape