DNA STRUCTURE AND FUNCTION Chapter 10

Identification of the Genetic Material Griffith’s Experiment Griffith’s Experiment: What made R strain into live S strain from dead one ? 협막(capsule)

그림 10.2

Infection of bacteriophage

Hershey-Chase Experiment 세균에 침투하는 것은 DNA 이며, 이것이 파지 자손을 증식시키는 유전물질임을 증명 (Sulfur 는 외피단백질의 원소가 되며, 인은 중핵 DNA 의 원소가 됨 )

표 10.1

A. DNA Structure DNA is a nucleic acid composed of nucleotide monomers. DNA nucleotide consists of: one phosphate group one deoxyribose sugar (5 carbon sugar) one nitrogenous base (G, A, C or T)

DNA is a double-stranded helix (Watson & Crick 1953). Sides of ladder make up sugar- phosphate “backbone”. Rungs (band) of ladder composed of base pairs joined by hydrogen bonds.

Pyrimidines (T & C) form hydrogen bonds with purines (A & G). Thymine pairs with Adenine, forming 2 hydrogen bonds. Cytosine pairs with Guanine, forming 3 hydrogen bonds.

DNA strands are antiparallel. 5’ to 3’ strand 3’ to 5’ strand Numbering of strands is based on position of deoxyribose sugars.

DNA is highly condensed. DNA is wrapped tightly around proteins(neucleosome) & folded. DNA must unwind for replication to occur.

B. DNA Replication Process by which DNA is duplicated. occurs during the S phase of Interphase is semiconservative (Meselson & Stahl) Exp. N15/N15 에서 부모세대 배양후 방사능 검사 N14 에서 1 세대 배양 ( 대장균 : 약 30 분 ) 후 방사능 검사 1 세대 세균을 N14 에서 2 세대 배양 후 방사능 검사 LH HH LLLH

Overview of DNA Replication:  Unreplicated DNA.  Strands “unzip” at several points creating replication forks.  Each strand serves as template for complementary nucleotides to H-bond.  New nucleotides of each daughter strand are linked.

Steps in DNA Replication:  Helicase breaks hydrogen bonds.  Binding proteins stabilize strands; prevent them from rejoining.  Primase makes an RNA primer.

 Free nucleotides move in & H-bond; DNA polymerase links nucleotides to each other starting at primer & working in the 5’ to 3’ direction.  DNA polymerase “proofreads” new strand (replaces incorrect bases).

 DNA replication is continuous on one strand.  DNA replication is discontinuous on other strand, producing Okazaki fragments.

 Repair enzymes remove RNA primers; Ligase connects Okazaki fragments. Ligase

Determine the base sequence of daughter DNA replicated from the following parental DNA strand. parental DNA C T A G G T A C T daughter DNA G A T C C A T G A

C. DNA Repair UV radiation damages DNA by causing thymine dimers to form. DNA damage can be repaired by photoreactivation or excision repair.

Photolyase is present and functional in prokaryotes, is present in lower eukaryotes (as yeast) where it is thought to have a minor role, and it has not been found in human cells.prokaryoteseukaryotes However, many higher eukaryotes, including humans, possess a homologous protein called cryptochrome that is involved in light-sensitive regulatory activities such as modulating circadian rhythms. homologouscryptochrome circadian rhythms Photolyases are flavoproteins and contain two light-harvesting cofactors. All photolyases contain the two-electron-reduced FADH 2 ; they are divided into two main classes based on the second cofactor, which may be either the pterin methenyltetrahydrofolate (MTHF) in folate photolyases or the deazaflavin 8-hydroxy-7,8-didemethyl-5-deazariboflavin (8-HDF) in deazaflavin photolyases.flavoproteins cofactors FADH 2pterindeazaflavin (from Wikipedia)

1. Photoreactivation – photolyase (enzyme) uses light energy to split dimer. 2. Excision repair - repair enzyme cuts out damaged area; DNA polymerase inserts replacement sequence & ligase seals backbone.

그림 10.12

3. Mismatch repair - enzymes proofread newly replicated DNA for base mispairing & correct the error. Faulty DNA repair results in chromosome breaks & an increased susceptibility to cancer. Ex. Xeroderma pigmentosum ( 열성유전 색소건피증 )

D. Comparison of DNA & RNA A = T G C 퓨린 : 피리미딘

오페론 : 세균에서의 유전자발현

E. Transcription Process by which a molecule of RNA is synthesized that is complementary to a specific sequence of DNA Occurs in the nucleus of eukaryotic cells & cytoplasm of prokaryotic cells. Is regulated by operons (bacterial cells) or transcription factors (multicellular organisms). Involves 3 stages: initiation, elongation & termination

1. Initiation RNA polymerase attaches to a promoter on DNA strand. Helicase unzips a short section of DNA. Free RNA nucleotides move in & H-bond to complementary bases on DNA template strand.

2. Elongation RNA polymerase links RNA nucleotides together in a 5’ to 3’ direction. Growing RNA strand peels away from DNA template. 3. Termination RNA polymerase detaches when it reaches a terminator. Completed RNA molecule is released from DNA template.

Usually, several copies of RNA are made at a time. Determine the base sequence of RNA transcribed from the following DNA template strand. DNA template C A G T A A G C C RNA strand G U C A U U C G G 123

Three major types of RNA are transcribed. mRNA (messenger RNA) - encodes genetic information from DNA & carries it into the cytoplasm. Each three consecutive mRNA bases forms a genetic code word (codon) that codes for a particular amino acid. codon 5’3’

rRNA (ribosomal RNA) - associates with proteins to form ribosomes. Subunits are separate in the cytoplasm, but join during protein synthesis (translation). large subunit small subunit

tRNA (transfer RNA) - transports specific amino acids to ribosome during protein synthesis (translation). Anticodon - specific sequence of 3 nucleotides; complementary to an mRNA codon. Amino acid accepting end Anticodon sequence determines the specific amino acid that binds to tRNA.

The Genetic Code

Eukaryotic mRNA must be processed before it exits nucleus & enters cytoplasm. nucleotide cap is added “poly A tail” is added (a series of adenines) introns are removed

F. Translation Process by which an mRNA sequence is translated into an amino acid sequence (polypeptide/protein). Occurs in the cytoplasm of eukaryotic & prokaryotic cells. Requires: mRNA, tRNAs, amino acids & ribosomes. Involves 3 stages: initiation, elongation & termination

The Genetic Code

1. Initiation Small ribosomal subunit binds to “start codon” [AUG] on mRNA molecule. AUG codon attracts initiator tRNA.

2. Elongation Large ribosomal subunit binds to small subunit. A second tRNA anticodon binds to the next mRNA codon. A peptide bond forms between the two amino acids.

Initiator tRNA is released. Ribosome moves down mRNA by 1 codon. A third tRNA anticodon binds to the next mRNA codon. A peptide bond forms between 2nd & 3rd amino acids.

tRNAs continue to add amino acids; polypeptide lengthens.

3. Termination Occurs when ribosome reaches an mRNA stop codon (UGA, UAG or UAA). Stop codons do NOT specify an amino acid. Last tRNA is released, ribosomal subunits separate & new polypeptide/protein is released.

Usually, several copies of the polypeptide/protein are made at a time. Some polypeptides must be altered before they can function

Determine the amino acid sequence a ribosome would translate from the following mRNA strand. mRNA C A U G G C U C A A U G A AlaGlnSTOPMet

Review: Genetic information flows in cell from DNA  RNA  protein. Each gene on DNA codes for production of a specific polypeptide/amino acid.

G. Mutation A physical change in the nucleotide sequence of DNA. May not affect phenotype (silent mutation). Can affect somatic cells (somatic mutation) or sex cells (germinal mutation). Can form spontaneously or be induced by a mutagen.

1. Point mutation - one DNA nucleotide replaces another missense mutation - point mutation that changes a codon to specify a different amino acid. Ex. sickle cell disease

nonsense mutation - point mutation that changes an amino acid-specifying codon into a stop codon. 2. Frameshift mutation - the insertion or deletion of DNA nucleotides; results in disruption of the reading frame. Ex. cystic fibrosis ( 낭포성 섬유증 ) 3. Expanding repeat - the # of copies of a 3 or 4 nucleotide sequence increases over several generations. Ex. myotonic dystrophy ( 근강직성 디스트로피증 )

표 11. 6

표 11. 5

Natural protection against mutation DNA proofreading DNA repair checking RNAs as they are made eliminating malformed proteins genetic code

Protection in genetic code: synonymous codons ( 동의 코돈 ) encode same amino acid, 3 rd position differs mutation in 2 nd codon position changes specification to similar amino acid mutation affects phenotype only if it alters protein’s function

p.201 그림 10B