I. DNA ReplicationA. Nucleotides added to each original template strand

1. added in 5’ to 3’ direction 2. The two strands of DNA are anti-parallel a. run in opposite directions

3. DNA polymerase a. Form a bond between - 3’ OH group of deoxyribose on last nucleotide - 5’ phosphate of the correct dNTP b. Add nucleotide and release diphosphate c. Move to next nucleotide on template

B. Initiation of DNA replication 1. Origins of replication a. Specific sequence of nucleotides b. recognized by proteins that bind to DNA - initiator proteins

2. DNA helix unwinds - caused by helicase enzymes

3. bacteria have a single origin of replication

B. Initiation of DNA replication 1. Origins of replication a. Specific sequence of nucleotides b. recognized by proteins that bind to DNA - initiator proteins

2. DNA helix unwinds - caused by helicase enzymes

3. bacteria have a single origin of replication

4. eukaryotes have multiple origins

a. form multiple replication bubbles - single stranded regions stabilized by single stranded binding proteins b. multiple replication forks

5. primase enzyme binds a. synthesizes short sequence of RNA b. DNA polymerase function requires RNA primer

~10 nucleotides long

c. primer sequence made on template - template - single strand of original DNA - complimentary bases

C. Elongation 1. elongation of complimentary strand – 5’ to 3’ a. done by DNA Polymerase - DNA polymerase can only add nucleotides to a pre-existing strand - requires RNA primer to begin - polymerase makes complimentary strand to template

2. Elimination of RNA Primer a. different DNA P’ase - eats away RNA with exonuclease * can degrade DNA from ends - adds in DNA - close gap in DNA - DNA Ligase

SHOW ANIMATION – Leading Strand

3. Bidirectional Elongation of new strands a. DNA replication proceeds only in 5’--> 3’ direction

b. Leading strand - polymerase adds nucleotides continuously 5’-->3’ from origin site - follows replication fork

c. Lagging strand - replication fork opening 3’ to 5’ – moving away from replication fork - DNA P’ase does NOT work 3’-5’

4. Elongation of lagging strand a. synthesized in short segments - Okazaki fragments

- each segment requires its own RNA primer - each fragment ~100-200 nucleotides long - each RNA primer replaced with DNA - fragments joined by DNA ligase enzyme

SHOW ANIMATION Lagging Strand

5. DNA editing during replication a. occasional errors by polymerase (1/10,000 bp)

b. DNA polymerase proof-reads

- checks each new nucleotide against template - if mismatched, it backs up and replaces it

D. Termination 1. Leading strand closes in on other lagging strand 2. Ligase completes new strand 3. Daughter strands paired with original strands - semi-conservative replication

E. Summary of Replication 1. Happens before cells divide - During Synthesis phase of cell cycle 2. Fueled by breaking phosphate bonds

3. Begins at origin sequences

4. Process determined by Replication enzymes

II. Transcription and TranslationA. Processes produce proteins from DNA using RNA 2. DNA a. nucleic acid polymer b. contains genes – segments of DNA that code for proteins c. Each gene codes for a different protein - one gene-one protein (enzyme) hypothesis

3. One gene-one enzyme hypothesis a. Each gene codes for a different enzyme

b. Metabolism occurs through pathways of chemical reactions c. Each step is catalyzed by a single enzyme

d. Study pathways by identifying metabolic needs of mutants - Beadle and Tatum studied metabolic pathway in mold

* some mutant molds require arginine* others require citrullene or ornithine

•hypothesis: each mutant lacks an enzyme

•See Figure 17.1

4. RNA a. nucleic acid polymer b. RNA is transcribed from DNA - messenger RNA - mRNA - RNA sequence determined by chemical rules

of base pairing - carries code from gene to protein

5. Protein – amino acid polymer a. Proteins are translated from RNA

b. genetic code uses triplets of bases to determine amino acid sequence

6. One way flow of information - DNA --> RNA --> Proteins

7. Location of activities in Eukaryotes a. Transcription of RNA from DNA occurs in cell nucleus

b. Translation of RNA into protein occurs in cytoplasm

8. In Prokaryotes there is no nucleus a. Translation occurs immediately after transcription

B. Transcription 1. Synthesis of mRNA 2. RNA nucleotides transcribed from DNA template 3. requires the enzyme RNA polymerase

4. RNA transcript made in 5’-3’ direction (like DNA) a. DNA template strand runs in 3’-5’ direction b. Corresponding DNA strand in 5’-3’direction is “coding strand”

5. Three stages to transcription process a. where to start - initiation b. transcribe whole message - elongation c. where to end - termination

C. Initiation 1. Promoter region a. sequence of DNA “upstream” from start point b. recognized by proteins

- proteins = transcription factors c. initial binding to TATA box - allows subsequent binding of RNA P’ase

d. Other transcription factors and RNA P’ase can bind

e. DNA unwinding beginsf. RNA polymerase begins synthesizing transcript

D. Elongation 1. RNA polymerase moves “down” DNA strand a. NTPs added to growing mRNA strand - proceeds in 5’ to 3’ direction b. single gene may be transcribed multiple times simultaneously - several RNA polymerase molecules can follow each other “down” the DNA template

E. Termination of Transcription 1. RNA polymerase reaches ‘terminator” sequence a. specific DNA sequences - RNA P’ase transcribes past termination sequence - mRNA strand released by polymerase - RNA polymerase released from DNA template

F. Genetic Code 1. 3 nucleotides code for each amino acid a. each triplet a “word” = codon - specifies a single amino acid

b. 20 amino acids total - 4 nucleotides to fill three codon positions - 43 combinations of nucleotides = 64 codons - more than one triplet can code for same amino acid

= redundancy

c. genetic code has start and stop signals - AUG = start - UAA, UAG, UGA= stop

3. Universality of the Genetic Code a. All organisms share almost identical code - occasional rare changes in few codons b. evidence for a single origin of all life - evolved billions of yrs ago - all existing organisms probably had a common ancestor

c. practical implications? - Can insert genes between different organisms - genetic engineering