DNA Structure: Key Concepts and Discoveries

DNA Structure

• DNA was discovered in 1953
• Scientists Francis Crick and James Watson determined the structure of DNA
• Linus Pauling and Maurice Wilkins were also involved in research
• Rosalind Franklin used X-ray crystallography to understand DNA structure
• Watson and Crick used Franklin's data and Chargaff's rules to determine the structure.
• Chargaff's rules showed that in DNA, the amounts of adenine and thymine were equal, and guanine and cytosine were equal.
• DNA is a double helix composed of two strands twisted around each other.
• Bases pair in a complementary manner: adenine with thymine, and guanine with cytosine.
• DNA is made up of four kinds of monomers called nucleotides
• Each nucleotide contains a deoxyribose sugar, a phosphate group, and a nitrogenous base
• Two types of nitrogenous bases are double-ringed purines (adenine and guanine)
• Two types of nitrogenous bases are single-ringed pyrimidines (cytosine and thymine).
• The phosphate group of one nucleotide joins covalently with the sugar molecule of the next.
• DNA is a long polymer of nucleotides with sugar-phosphate backbones, and nucleotide bases that stick out.
• The sugar-phosphate groups form a "backbone" for each strand.
• Carbon atoms of the five-carbon sugar are numbered clockwise.
• The phosphate group attaches to the 5' carbon of one nucleotide and 3' carbon of the next.
• Complementary strands are held together by hydrogen bonds between the bases.

DNA Arrangement in Cells

• Prokaryotic DNA is simpler than eukaryotic DNA
• Most prokaryotes have one circular chromosome in an area called the nucleoid.
• Eukaryotic DNA is bound to proteins known as histones that form nucleosomes.
• Nucleosomes stack compactly onto each other to form a 30-nm-wide fiber.
• This fiber is further coiled into a more compact structure during the metaphase stage of mitosis.
• Eukaryotic chromosomes have two distinct regions
• Darkly staining regions usually contain inactive genes
• Lightly staining regions usually contain active genes.

DNA Replication

• DNA is copied during DNA replication in the synthesis phase (S phase) of cell cycle before cell division.
• The double helix separates into two strands.
• Each strand serves as a template from which a new complementary strand.
• Semiconservative Replication: Each new double strand is composed of one parental strand and one new daughter strand.

DNA Replication in Eukaryotes

• Eukaryotic DNA is very complex and involves several enzymes and proteins for replication.
• Replication starts at specific nucleotide sequences called origins of replication and proceeds in both directions.
• Replication forks (Y-shapes structures) are formed at the origins of replication as the DNA double helix opens up.
• DNA polymerase adds DNA nucleotides to the 3' end of the template.
• Leading strand is synthesized continuously toward the replication fork, and this allows the DNA polymerase to add DNA nucleotides continuously.
• Lagging strand is synthesized in short fragments called Okazaki fragments, this means that DNA is not synthesized continuously in the lagging strand.
• RNA primers are required to start the synthesis of Okazaki fragments.
• RNA primers are removed from the lagging strand by DNA polymerase and replaced by DNA nucleotides.
• The gaps between fragments are sealed by DNA ligase.

DNA Replication in Prokaryotes

• Easier DNA replication process due to the small genome size relative to eukaryotic DNA.
• Replication starts at a single origin and proceeds in both directions around the circular chromosome
• There is rapid replication rate of roughly 1000 nucleotides/second.

DNA Repair

• DNA polymerase proofreads incoming bases after adding to prevent errors.
• Mismatch repair mechanisms detect and remove incorrect bases later in the process.
• Nucleotide excision repair removes damaged or incorrect bases and nucleotides.
• Errors lead to mutations (permanent DNA sequence changes), some of which can result in diseases like cancer.

Transcription

• DNA is transcribed into mRNA via transcription and the processes lead to protein creation within the cell.
• DNA serves to provide the genetic information required for the formation of proteins necessary for the functions of a cell.
• Transcription involves the synthesis of an mRNA molecule that is complementary to the DNA template strand.
• Proceeds through initiation, elongation, and termination stages in Eukaryotes and Prokaryotes
• Transcription takes place in the nucleus in eukaryotic cells and the cytoplasm in prokaryotic cells.
• Multiple RNA polymerase enzymes are used in transcribing different genes .
• DNA is partially unwound to produce a transcription bubble during initiation.
• Promoter is the region where transcription factors and RNA polymerase bind to initiate transcription.
• RNA polymerase proceeds along the template strand, synthesizing the mRNA.
• RNA polymerase continues the process while unwinding and rewinding the DNA.
• Terminator signals tell the RNA polymerase to stop and release the newly formed mRNA in prokaryotic cell termination.

RNA Processing in Eukaryotes

• Eukaryotic mRNAs undergo processing steps before leaving the nucleus.
• Modifications protect mRNA from degradation and aid in protein synthesis.
• A 5' cap and poly-A tail are added to the 5' and 3' ends, respectively.
• Introns (non-coding regions) are removed by splicing
• Exons (coding regions) are joined together.
• The mRNA is transported to cytoplasm after processing

Translation

• Translation involves using the mRNA code to synthesize proteins.
• It uses ribosomes, tRNA molecules, and amino acids.
• mRNA specifies the sequence of amino acids that should be linked to form a protein.
• Ribosomes are composed of rRNA and multiple proteins and function to help facilitate translation of process.
• tRNAs bring amino acids to the ribosome based on the mRNA codons.
• Several steps: initiation, elongation, and termination
• Initiation: Brings together the small subunit of the ribosome, mRNA molecule, tRNA molecule and initiation factors
• Elongation: Amino acids are added to the growing polypeptide chain.
• Termination: The stop codon is recognized and the translation machinery leaves the template and disassembles.
• The process generates a polypeptide chain which then folds into a functional protein.

Gene Regulation

• Gene expression is controlled in prokaryotes mainly at the transcriptional level (how often a gene is transcribed).
• Prokaryotes use operons (e.g., lac operon). The operons can be controlled by repressors that bind to DNA and block transcription or activators that do the opposite.
• Gene expression in eukaryotes is regulated at multiple levels, including epigenetic, transcriptional, post-transcriptional, translational, and post-translational levels.
• Eukaryotic cells use a complex system of proteins to control gene expression. These proteins influence where and when genes or certain parts of genes are expressed in response to signals.

Alternative RNA Splicing

• Eukaryotic genes have multiple different sites that can be used during splicing
• Splicing mechanism can sometimes fail to identify intron boundaries which can affect the proteins produced from genes.
• Alternative splicing generates different protein isoforms from the same gene, influencing cell specialization