Things to do today? • Double Helix Essay - bring to lab! • Write your own exam question. • Review material (including today) for Friday’s exam.

Exam 2: Friday, March 15th.

Exam 2 (3/15) will include a metabolic pathways handout Handout Copy is on Web Site

buildingdashboard.net/brown A competition between dorms to reduce their energy usage by the most percentage compared to the month prior to the competition. Prizes are a dinner with President Paxson, bikes, and other prizes. Only those who are registered can have the opportunity to win the prizes. buildingdashboard.net/brown

DNA Replication All polymerases, including DNA polymerase, add bases to the 3’ end of a growing strand. Direction of Fork Movement 5’ 3’ Replication is not as simple as it once seemed! DNA replication is Continuous on one strand: And Discontinuous on the other: The 3’ and 5’ labels on this slide have been corrected!

Every piece of DNA begins with a short strand of RNA: 3’ Every piece of DNA begins with a short strand of RNA: 5’ Primase makes an RNA primer Direction of Fork Movement 3’ 5’ 5’ The 3’ and 5’ labels on this slide have been corrected!

Replication continues until the RNA fragments are replaced by DNA 3’ Replication continues until the RNA fragments are replaced by DNA 5’ DNA Ligase joins fragments together Direction of Fork Movement 3’ 5’ The 3’ and 5’ labels on this slide have been corrected!

But it’s even more complex than that — there are at least 2 distinct DNA polymerases on each replication fork, and a DNA-unwinding protein called DNA helicase

Freeman 4/e (p. 267)

Transcription & Translation The “Central Dogma” Transcription RNA polymerase Promoters Introns & Exons The Genetic Code The Ribosome tRNA, rRNA, and mRNA Translation

The“Central Dogma”

DNA RNA Protein The“Central Dogma” replication transcription translation Protein

RNA: 1) Ribose sugar 2) Uracil in Place of Thymine

RNA is synthesized against a DNA template by RNA polymerase, a complex multisubunit enzyme.

DNA RNA Movement of RNA polymerase along DNA can actually be visualized in the EM:

Base-pairing during transcription follows the Watson-Crick pattern. 3’ 5’ 5’ 3’ RNA polymerase (like DNA polymerase) produces a new strand in the 5’ to 3’ direction. As a result, RNA is complementary to one strand of DNA (the “template”) and nearly identical to the other (the “coding strand”)

How does RNA polymerase know “which” strand to copy? DNA sequences known as “promoters” serve as RNA polymerase binding sites. And RNA polymerase binds to a promoter oriented in a particular direction. So, either strand may be transcribed, depending on location and direction of Promoter sequences.

Promoters have general features that (sometimes) enable them to be recognized in a DNA sequence.

mRNA (messenger RNA) tRNA (transfer RNA) rRNA (ribosomal RNA) DNA mRNA (messenger RNA) tRNA (transfer RNA) rRNA (ribosomal RNA) Transcription produces three general classes* of RNA, each of which plays a role in translation (protein synthesis) * actually, there are many more classes of small RNA molecules that perform important functions in the cell, including gene regulation and RNA splicing.

Sometimes it did. But sometimes it didn’t! He thought mRNA would match up perfectly to the DNA from which it was transcribed: In 1975, when Phillip Sharp first compared mRNA sequences to the DNA template strands that produced them, he was surprised. Sometimes it did. But sometimes it didn’t!

Exons = Expressed sequences. Introns = Intervening sequences In eukaryotes (but not in prokaryotes), most mRNAs are made in a pre-mRNA form, which is then cut and spliced to remove intervening sequences (“introns”) before leaving the nucleus. Exons = Expressed sequences. Introns = Intervening sequences

Next question: How does the sequence of bases in RNA determine the sequence of amino acids in a polypeptide? ??

George Gamow (physicist & SciFi writer) 1) There are 20 common amino acids 2) The code must specify 20 different possibilities in each "word" 3) If one nucleotide corresponded to one amino acid, only 4 words could be formed (only 4 bases in RNA). 4) If two bases specified an AA, that would only allow 16 (4x4) words. Not enough. 5) But three bases per AA would work (4x4x4=64, more than enough to produce the minimum of 20 words needed). 6) Since nature always works in the most economical way, the genetic code must be written in triplets - three bases at a time.

Translation begins with a “start” codon, usually AUG... UAA 3’ Translation begins with a “start” codon, usually AUG... … and ends with 1 of 3 “stop” codons.

But Translation requires a complex “machine” to make it all work: Ribosomes are the sites of protein synthesis Ribosomes bind to the beginning of a mRNA molecule (the AUG initiator near the 5’ end), and catalyze the polymerization of amino acids to form a polypeptide.

Ribosomes (250Å diameter) seen in electron microscope. 3-D functional model of ribosome prepared from EM images (Joachim Frank, SUNY Albany):

Ribosomes consist of two subunits (large & small) Ribosomes are composed of RNA and protein. (Eukaryotic: 4 rRNA molecules and 83 proteins)

The detailed structure of the ribosome is now known at the atomic level. rRNA forms the functional core of the ribosome.

Q: How does the ribosome pick the “right” amino acid to match each codon in the message? UAC met CAC his 5’ AUGCACGGAUUUUCCAAC . . . . . . . UAA 3’ A: It doesn’t! Amino acids are bound to transfer RNA (tRNA) molecules, which base-pair to mRNA codons.

tRNAs are short (75-90 bases) RNA molecules tRNAs are short (75-90 bases) RNA molecules. Enzymes attach the appropriate Amino Acid to each tRNA based on the tRNA’s own base sequence.