BMSC 210 - Microbiology: DNA and RNA Structure, Function, and Replication

Discovery and Characterization of DNA

• Historical Background: DNA was discovered in the late 1800s but was not recognized as the hereditary material until the mid-20th century.
• Mendel's Contributions: Gregor Mendel documented patterns of inheritance in pea plants; laid the foundation for genetics through the study of traits and heredity.
• Chromosomal Theory of Inheritance: Proposed by Sutton and Boveri, indicating that chromosomes are the carriers of genetic material.
• One Gene - One Polypeptide Theory: Beadle and Tatum's hypothesis that each gene encodes the production of a specific polypeptide.

Griffith’s Transformation Experiments

• Experiment Setup: Demonstrated that non-pathogenic bacteria could transform into pathogenic forms when exposed to heat-killed pathogenic strains.
• Key Findings: Transformation suggested a “transforming principle” responsible for hereditary material.

Avery, MacLeod and McCarty Experiment

• Objective: Identified DNA as the molecule responsible for transformation.
• Experiments Conducted:
  • Control: Used rough strain (nonvirulent).
  • Experiment 1: Heat-killed smooth strain (virulent) injected into mice led to death.
  • Experiment 2: Mixed heat-killed smooth strain with rough strain; transformation occurred.
  • Experiment 3: Enzymes that degrade proteins, RNA, and DNA were utilized; only when DNA was degraded, transformation did not occur.
• Conclusion: Transformation requires DNA; proved DNA is the genetic material.

Hershey and Chase Experiment

• Objective: To determine whether DNA or protein coat of phage was the genetic material entering host bacteria.
• Methodology:
  • The use of 32P to label DNA and 35S to label proteins.
  • Infected bacteria with labeled phages and determined which label was present in the offspring phages.
• Results: Only 32P-labeled DNA entered the bacterial cells, confirming that DNA is the genetic material.

Key Events Timeline

• 1869: Miescher identifies DNA.
• 1902: Sutton proposes chromosome theory.
• 1928: Griffith discovers transformation.
• 1944: Avery, MacLeod, and McCarty prove DNA is the transformative agent.
• 1952: Hershey and Chase confirm DNA is genetic material.
• 1953: Watson and Crick propose the double helix structure of DNA.

Structure of DNA

• Deoxynucleotides: Building blocks are dNTP, consisting of a deoxyribose sugar, phosphate group, and nitrogenous base.
• Double Helix:
  • Found in two antiparallel strands.
  • Complementary base pairing (A-T; C-G).
  • Features major and minor grooves.
• Replication Characteristics: Synthesized in the 5' to 3' direction with a leading and lagging strand during replication.

Function of DNA

• Genetic Information Storage: Serves as the template for genetic instructions necessary for cellular functions.
• Organizational Structure: Contained within chromosomes; variations between prokaryotes and eukaryotes.
• Types of DNA:
  • Prokaryotes: Generally single, circular DNA.
  • Eukaryotes: Linear chromosomes.

Central Dogma of Molecular Biology

• Flow of Information: Understanding how DNA is transcribed into RNA and translated into proteins is crucial.
• Replication: DNA replicates semi-conservatively; each new DNA molecule contains one old and one new strand.
• Transcription: Process of synthesizing RNA from DNA.
  • RNA polymerase binds to the DNA template and synthesizes mRNA in the 5' to 3' direction.
• Translation: Involves ribosomes synthesizing proteins based on codons in mRNA, mediated by tRNA.

DNA Replication Enzymes

• Helicase: Unwinds the DNA helix.
• DNA Polymerase III: Main enzyme synthesizing new DNA strands.
• Ligase: Seals nicks between Okazaki fragments on the lagging strand.
• Primase: Synthesizes RNA primers necessary for DNA synthesis.
• Topoisomerase: Relieves strain during DNA unwinding.

Structure and Function of RNA

• Types of RNA:
  • mRNA: Carries genetic information from DNA for protein synthesis.
  • tRNA: Transfers amino acids during protein synthesis.
  • rRNA: Constitutes the ribosome's structural and functional components, catalyzing peptide bond formation.

Transcription Steps**

• Initiation: RNA polymerase binds to the promoter region, unwinding DNA.
• Elongation: RNA is synthesized as RNA polymerase transcribes the template DNA strand into mRNA.
• Termination: Transcription terminates at specific signals (e.g., stem-loop structures).

Translation Process**

• Ribosome: Site for translation; consists of large and small subunits.
• Initiation: Begins with recognition of the start codon by tRNA.
• Elongation: Amino acids are sequentially added to form a growing polypeptide chain.
• Termination: Reached upon encountering a stop codon, releasing the newly synthesized protein.

Comparison of Prokaryotic and Eukaryotic Translation

• Location: In prokaryotes, translation occurs concurrently with transcription; in eukaryotes, translation occurs in the cytoplasm after RNA processing.
• Ribosome Size: Prokaryotic ribosomes are 70S; eukaryotic ribosomes are 80S.
• Initiator tRNA: Prokaryotes use fMet-tRNA, while eukaryotes use Met-tRNA.

Protein Processing and Folding**

• Newly synthesized polypeptides typically undergo folding and post-translational modifications to become functional proteins. Chaperones assist in proper folding, and energy is required for efficient processing.