UNIT Pearson Education, Inc.

Transcription

1 UNI 2. Mendel and the ene Idea (h11) B. hromosomal Basis of Inheritance (h12). Molecular Basis of Inheritance (h13) D. ene Expression from ene to Protein (h 14) Where are we going? H13 More discoveries concerning the heritable material! What is it? What might its structure be? Replication and how it works Listen to an interview of Watson We will try to keep our eye on big picture as we look over details of replication!

2 Figure 13.21b hromatid (700 nm) 30-nm fiber Loops Scaffold 300-nm fiber Reminder of structure! Replicated chromosome (1,400 nm)

3 Figure 13.21a DN double helix (2 nm in diameter) Nucleosome (10 nm in diameter) Histones Histone tail H1 Histones are a protein

4 Figure DN double helix (2 nm in diameter) Nucleosome (10 nm in diameter) 30-nm fiber hromatid (700 nm) Loops Scaffold Histones Histone tail H1 300-nm fiber Replicated chromosome (1,400 nm)

5 Remember Morgan sex linked traits.. He confirmed that genes are on chromosomes But chromosomes are made up of protein ND DN which one is doing the important job of harboring information? We will focus on these 3 classic experiments highlighted in this chapter riffith (Fred) Hershey and hase (lfred and Martha) Meselson and Stahl (Matt and Frank)

6 Early 1900s everyone was focusing on proteins as hereditary material DN was big but monotonous seemed to have no specific cellular function Proteins were enzymes! nd had structural jobs in cells! Proteins were also known to be polymers of numerous amino acids. his 20 amino acid "alphabet of proteins could be arranged into more unique information-carrying structures than the four-letter alphabet of DN

7 riffith (rying to develop a vaccine) urned to a bacterial pathogen Streptococcus pneumoniae Is it gram positive or negative? %22_1936.jpg

8 his species (Streptococcus pneumoniae) is one of the species that can cause meningitis (along with Neisseria meningitidis). What is meningitis? Did you get your meningitis vaccine? Note you can also get viral meningitis but usually not as dangerous-no vaccine. Streptococcus pneumoniae is also a cause of pneumonia along with a bunch of other organisms What is pneumonia?

9

10 So what did he do?? 2 strains what do we mean by strains? One virulent and not so virulent (What do we mean by virulent?)

11 Figure 13.2 Experiment Living S cells Living R cells Heat-killed S cells Mixture of heat-killed S cells and living R cells Results Mouse dies Mouse healthy Mouse healthy Mouse dies Work by very identified the transforming substance as DN (but others not convinced) Living S cells

12 ransformation-did not really understand mechanism an we do this-pick up DN from our environment? Why does this ability freak us out.what do we feed chickens pigs and cows to make them grow fast? Where do we put their manure waste?? Where can this manure waste go?

13 Hershey and hase (1952) (lfred and Martha) heir work also pointed to DN rather than proteins Bacteriophages what are they???? (worked with one called 2) E. coli bacterial cell-who is E. coli? t this time they knew that viruses could infect bacterial cells with heritable material and...that material that is put into bacterial cells hijacks or reprograms the cell to makes new viruses. Is it the coat (protein) or is it the stuff inside (DN)? Which is actually entering the cell and reprogramming it?

14 Some phages grown in media for a couple hrs with radioactive Sulphur which should be incorporated into some coat proteins (Methionine, ysteine) Other phages grown in media for a couple hrs with radioactive Phosphorus.which should be incorporated into DN

15 hese phages were allowed to infect bacteria. Knocked off phage from cell and then centrifuged to concentrate cells. Where is Phosphorus (DN)? Where is Sulfur (Protein oat)?? Batch with P (DN) labeled, found in cells. Batch with S (Protein oat) labeled, found in liquid Was the protein coat passed on? hus, the Hershey hase experiment helped confirm that DN, not protein, is the genetic material.

16 Figure 13.4a Experiment Batch 1: Radioactive sulfur ( 35 S) in phage protein 1 Labeled phages 2 gitation frees 3 infect cells. outside phage parts from cells. Radioactive protein entrifuged cells form a pellet. 4 Radioactivity (phage protein) found in liquid entrifuge Pellet

17 Figure 13.4b Experiment Batch 2: Radioactive phosphorus ( 32 P) in phage DN 1 Labeled phages 2 gitation frees 3 infect cells. outside phage parts from cells. Radioactive DN entrifuged cells form a pellet. entrifuge Pellet 4 Radioactivity (phage DN) found in pellet

18 Watson-rick Model predicted. Each of two daughter molecules would have one parental strand and one newly made!

19 he annual "degree granting" party of Max Delbrück's phage group held in the house shared by Meselson and Stahl at altech. (L-R) Harry Rubin, Max Delbrück, Rene ohen, Matt Meselson, Frank Stahl

20 Meselson and Stahl-clever experiment What did they do?? SBLE ISOOPES! rew up some E. coli in a medium with heavy N (or 15 N)-incorporated into DN bases (,,,). ransferred to a light N (or 14 N) medium and let E. coli replicate there. hen start taking subsamples of this mix over time. an separate by weight using centrifuge get bands or layers sked what N (heavy or light) is going to be in this population of E. coli at different times?

21 Figure 13.11a Experiment 1 Bacteria cultured in medium with 15 N (heavy isotope) 2 Bacteria transferred to medium with 14 N (lighter isotope) Results DN sample centrifuged after first replication 3 4 DN sample centrifuged after second replication Less dense More dense

22 Figure First Second Move from heavy into Parent light cell N media-what replication replication will happen (a) onservative when you model centrifuge? (b) Semiconservative model (c) Dispersive model Dark blue=heavy 15N

23 Figure Second Replication. (a) onservative model How many bands? Parent cell First replication Second replication (b) Semiconservative model (c) Dispersive model Dark blue=heavy 15N

24 Figure 13.1 Watson and rick james_watson_on_how_he_discovered_dna.html

25 Watson interview! What did you think was interesting? What was unusual? Surprising? Insights into how science works?

26 First some info about structure of DN!

27 Figure 13.7 end Hydrogen bond end 3.4 nm 1 nm 0.34 nm end end (a) Key features of DN structure (b) Partial chemical structure (c) Space-filling model

28 Figure 13.7a 3.4 nm 1 nm 0.34 nm (a) Key features of DN structure

29 Figure 13.7b Phosphate group What are the blue pentagons? end 1 2 Hydrogen bond end Nitrogen rich bases,,, and have double rings -two double rings would not fit so bonds with and with. end nucleotide is a phosphate molecule + a sugar molecule + a nitrogen rich base end

30 Figure 13.5 Sugar phosphate backbone end Nitrogenous bases hymine () denine () ytosine () end DN nucleotide uanine ()

31 Figure New strand emplate strand Sugar Phosphate Base DN polymerase Nucleotide P i P Pyrophosphate 2 P i

32 Figure (a) Parental molecule (b) Separation of parental strands into templates (c) Formation of new strands complementary to template strands through addition of nucleotides

33 DN replication Where does it take place? When does it take place? How does it work??

34 Origin of replication? Replication fork? 3 5? (skip) RN primer (made by pink blob=primase). Other blobs (green=helicase=unzips) (blue-=topoisomerase) (gray=single stranded binding proteins) Replication fork RN primer Figure 13.12

35 Figure DN polymerase III (salmon blob) Leading strand Overview Origin of replication Lagging strand Primer Binds then walks along, adding complementary Leading nucleotide bases (,, and ) Lagging to strand the strand of DN strand (in the 5 to 3 direction-but you do not need to know this). It is making a leading strand! Origin of replication RN primer Parental DN DN pol III ontinuous elongation in the to direction

36 Figure 13.16a Lets zoom in to #1 and look at the lagging strand. Leading strand Overview Lagging Origin of replication strand Lagging strand Overall directions of replication Leading strand

37 Figure 13.16b-1 1 RN primer emplate strand

38 Figure 13.16b-2 1 RN primer. emplate strand RN primer for fragment 1 2 DN pol III makes Okazaki fragment 1. Once again DN polymerase III binds and then walks along adding new complementary nucleotide bases (,, and ) to the strand of DN.

39 1 RN primer. emplate strand RN primer for fragment 1 2 DN pol III makes Okazaki fragment 1. 3 DN pol III detaches. Okazaki fragment 1 What happened? Why did it stop? What is fragment called? Where is DN pol III going to go next??

40 Figure 13.16c-1 RN primer for fragment 2 Okazaki fragment 2 4 DN pol III makes Okazaki fragment 2. Now you have all these bits what has to happen next? nd who does that? an you leave the RN primer pieces in there?

41 Figure 13.16c-2 RN primer for fragment 2 Okazaki fragment 2 4 DN pol III makes Okazaki fragment 2. 5 DN pol I replaces RN with DN. different DN polymerase comes in to replace them! (I vs III)

42 Figure Leading strand template Overview Origin of replication Leading strand Lagging strand Single-strand binding proteins Helicase Parental DN Lagging strand template DN pol III Primer Primase Leading strand DN pol III Lagging strand Leading strand Overall directions of replication Lagging strand DN pol I DN ligase

43 Figure New strand emplate strand Sugar Phosphate Base DN polymerase Nucleotide Which DN polymerase is this? P i P Pyrophosphate

44 he diagram below shows a replication bubble with synthesis of the leading and lagging strands on both sides of the bubble. he parental DN is shown in dark blue, the newly synthesized DN is light blue, and the RN primers associated with each strand are red. he origin of replication is indicated by the black dots on the parental strands. Rank the primers (the red specks) in the order they were produced. If two primers were produced at the same time, overlap them.

45 he diagram below shows a replication bubble with synthesis of the leading and lagging strands on both sides of the bubble. he parental DN is shown in dark blue, the newly synthesized DN is light blue, and the RN primers associated with each strand are red. he origin of replication is indicated by the black dots on the parental strands. Rank the primers in the order they were produced. If two primers were produced at the same time, overlap them. a and h then b and g then c and f and finally e and d

46 rue of Leading strand, Lagging strand, or Both???? Daughter strand elongates away from replication fork Multiple primers needed Made in segments Made continuously Daughter strand elongates toward replication fork

47 rue of Leading strand, Lagging strand, or Both???? Daughter strand elongates away from replication fork Lag Multiple primers needed Lag Made in segments Lag Made continuously Lead Daughter strand elongates toward replication fork Lead

48 In an analysis of the nucleotide composition of DN, which of the following will be found? (Imagine counting number of nucleotides of each type) = and = + = + = + = +

49 In an analysis of the nucleotide composition of DN, which of the following will be found? = and = + = + = + = +

50 ytosine makes up 42% of the nucleotides in a sample of DN from an organism. pproximately what percentage of the nucleotides in this sample will be thymine? 31% 42% 8% 16% It cannot be determined from the information provided.

51 ytosine makes up 42% of the nucleotides in a sample of DN from an organism. pproximately what percentage of the nucleotides in this sample will be thymine? 31% 42% 8% 16% It cannot be determined from the information provided.