Homework

1. Homework Read Chapter 1 of ECB Due week from this Wednesday. (extra time for books to come) Homework Set #1 on web Due at start of class next Monday Due to your requests: Lecture on : Evolution Protein folding

2. What is Physics 498Bio? Biophysics or Biological Physics? Mathematician Stanislaw Ulam: “Ask not what physics can do for biology, ask what biology can do for physics. The first is biophysics, the second biological physics. Biological physics is a subset of physics. Wolfgang Pauli: “The history of physics is a history of making concepts.” The idea in biological physics is to untangle the incredibly complex of biology in terms of so-called simple ideas of physics. Eventually, one gets a greater understanding and new technology. Biological physics : under the mantle of complex physics, where the complexity comes from biology. But it can’t be too complicated, otherwise you never get anywhere. At present, to deal with biological systems in their living environment is much too complex. Eventually, it might not be so, but for now it is. Protein Folding—good example of Biological Physics

3. Biological Physics“Good” Problem: Protein Folding Unfolded  Folded Hans Frauenfelder , founder of biological physics. Inactive  Active Wolynes, PNAS, 1998 Energy/ Entropy Landscape of Protein Folding (More later…)

4. Biophysics vs. Biological Physics We’ll studying both, but emphasis on biophysics – get new biology by applying new physics…lasers, x-rays, microscopes

5. The Language of Life “I know it when I see it.” -- Justice Potter Steward in a 1964 Supreme Court case on pornography What things are alive? Qualities we associate with living being: —it moves, it reproduces, it eats. prerequisite for living Trees don’t move, A person can be alive even if they are unable to move. People are certainly alive if they’re born sterile, or become so. Viruses can’t reproduce by themselves —they high-jack apparatus of their bacterial hosts to do the job. Is a virus alive? People disagree. Eating –yes, although the must include trees: drinking (water) and taking in energy via sunlight.

6. Living defined at the molecular level At the molecular level, the definition, surprisingly, becomes somewhat less complex. All living organism consist of complex, heterogeneous macromolecules Necessary, but not sufficient. A bag of starch, mixed in with other polymers, is clearly not alive—but is does appear to be necessary. All living organisms consist and make complex, heterogeneous macromolecules. They do this by ingesting them and making them from simple compounds. Amazingly, through evolution, organism have survived by making only 4 macromolecules

7. What do you make? The 4 essential macromolecules Each macromolecules, made from small pieces.(A) DNA made up of nucleotides, (B) Proteins, made up of amino acids; (C) Lipid molecule from fatty acids (generally hydrophobic), often in cell membrane, (D) Branch carbohydrate from the simple sugars. Much of discussion from “Physical Biology of the Cell”, by R. Phillips, Jane Kondev, and Julie Theriot Protein (Branched) carbohydrate Lipid (cell membrane) (Fat) DNA

8. Two Great Polymer Languages(via Francis Crick) Nucleic Acid alphabet = 4 nucleotides Proteins alphabet = 20 amino acids

9. Few # of alphabets diverse polymers Length of a polymer can vary enormously--from a single one to 100 million or so --hence the number of possible combinations that make up the polymer, is enormous. In biology, polymer –106-108 monomers long. Let’s say N equals 15--a very short polymer. So for this short polymer, and alphabet = 4, that’s 415 = 230 = 1,073,741,824 or over a billion different combinations. Adding more complexity : Isomers Two Polymers : made up of the exactly the same type of monomers and have the same order of monomers, there can be an enormous variation in their physical placement, known as isomers. An isomer is where you have the same number of atoms in the two polymers, but the arrangements of these atoms are different. For example, each monomer can be connected to another monomer either straight or bent fashion. Then there is 2n4n = 8n = 23n= 245 =35,184,372,088,832 or over 35 trillion different polymers of length N=15. So, from a relatively simple set, one can get tremendous diversity.

10. Get to know your neighbors You will have to report to the whole class immediately afterwards! –so listen up! With a partner (who you don’t know)… Tell your name, your year (undergrad, vs. grad.) What you want to be when you “grow up” Tell one thing that’s surprising about yourself. An example (me): This is Paul. He’s a senior citizen. (Ph.D. 1990, Physics, but really Biophysics) When he grows up (in the next stage of his life), I want to be a ski instructor. Something surprising about myself: 6 years ago I had an unfortunate incident while in San Diego hitting a car head-on while riding my bicycle. After months of hospitalization and rehab, I can do most things, but have trouble with my leg and arm. Been arrested multiple times.

11. DNA is made up of A,T,C,G in a double helix of anti-parallel strands 3.4 Å 3.4 nm per ~10 base-pairs = 1 turn (360º) Must come apart for bases to be read. Anti-parallel has interesting interpretation for way DNA is made. One enzyme that read 5’ 3’ or two, with another reading 3’ 5’?

12. Structure of DNA A-T, G-C bond, line up Two strands are oppositely oriented Form a minor and major groove U RNA U 6 Å 12 Å Every 3 base pairs encode for one amino acid

13. DNA: series of 4 nucleotides (bases): A,T,G,C Transcription [DNA & RNA similar]  RNA: series of 4 nucleotides (bases): A,U,G,C  Translation [RNA & Proteins different] http://learn.genetics.utah.edu/units/basics/transcribe/ DNA RNA ProteinsCentral Dogma of Biology Proteins: series of 20 amino acids: Met-Ala-Val-… each coded by 3 bases  amino acid AUG Methionine; GCU  Alanine; GUU Valine Proteins are 3-D strings of linear amino acids Do everything: structure, enzymes…

14. DNA  RNA  Amino acids Transcribe translate (1 codon = 3 bases) Genetic code U is a slightly altered T Central Dogma of Biology

15. Minimal knowledge about Nucleotides • 4 nucleotides: A,T,G,C • A=T ≈ 2kT two hydrogen bonds G=C ≈ 4kT three hydrogen bonds • Many weak bonds…very strong overall structure. DNA is stable. • Requires enzyme/ATP to split apart, to do its thing: replicate, transcribe. • ATP– universal food source of all cells. ≈ 25 kT ≈ 100pN-nm.

16. DNA: A-T 2 H bonds; G-C 3 H bonds Thymine Cytosine Hydrogen Bonds (2kT) Adenine Guanine G-Cmore stable than A-T • – p stacking keeps it together (Grease); Phosphate negative charge makes it water soluble (Sort of like soap)

17. E1 E0 Temp, T Boltzman factor + Partition function(review of basic Stat. Mech. – see Kittel, Thermal Physics If T = 0 ºK, what proportion of particles will be in E1, Eo? Answer: pop(Eo) = 1pop(E1) = 0 If T > 0 ºK, what proportion of particles will be in E1, Eo? e-(E1-Eo)/kBT J= represents jth state Z = partition function

18. Simple case: Ball in gravitational field. Thermal fluctuations, finite probability of being at height, h. E = ?? Partition Function for 2-state system As ball gets smaller, probability gets smaller / larger ? “Ball” the size of O2? Why can you breathe standing up? What is 1/e height for O2? For O2, 1/e height is ~10 km ~height of Mt. Everest. (10 km is “death zone”) Probability of dying if you go over 20,000 ft is 10% for every trip!!

19. Two states A – B bonded: E ~ -5 kT(a few H-bonds) A , B not bonded: E = ?? A + B A – B 0   149 molecules 148 will be A - B 1 will be A, B

20. DNA double helix: Many weak (H-bonds), makes for very stable structure. If you have many weak bonds (e.g. each bond only few kT) you can get a biomolecule that will not fall apart. H bonded ~ 2 kT Zipped vs. unzipped What if just one bond? Bond/unbound? What if 10 weak bonds? Many base pairs, essentially completely stable. Still have end-fraying, but probability that whole thing comes apart– essentially zero. [Need enzymes to separate.] With proteins, lots of hydrogen and weak bonds – have conformational dynamics, but rarely fall apart!

21. Evaluate class 1. What was the most interesting thing you learned in class today? 2. What are you confused about? 3. Related to today’s subject, what would you like to know more about? 4. Any helpful comments. Put your name in upper right-corner. Then tear off your name before turning in. (That way you can be brutally honest!) Answer, and turn in at the end of class. (I’ll give you ~5 minutes.)

22. Cyrus Eric Josh Matthias Pengfei Wylie Charles Alireza Thuy (Vietnamese) Kiran Xin Anthony Pavel Vishel Anne (my TA)

24. Size Scales of DNA (+ Protein) Chromatin = Complex of DNA + Protein (histones + non-histones) Nucleotides [4 Diff. types, A,T,C,G] 8/17/06 3 x 109 = 3 billion ~ 1 meter Flexibility of DNA? ~ 1 meter packed in 3-10 mm (size of nucleus) # chromosomes? 46 (ca. 50) Length/chromosomes? ~ 1/50 meter = 2 cm!

25. Cell Size Bacteria - 1 mm Eukaryotic cell – 10-100 mm 10-100 mm  10-30 mm 1 mm (Nucleus 3-10 mm) How much DNA inside of every single cell? 1 meter So a meter of DNA must pack 3-10 mm! What does this tell about bendability of DNA? Like spaghetti, uncooked or cooked? See how this is measured using magnetic tweezers Interesting factoids: 1.  1014 cells in body… …more stars than in Milky Way Galaxy. 2.  200 different types of cells in body.

26. You are what you eat  Cleave get 100 pN-nm Energy (food) Energy (body can use) = Adenosine triphosphate, ATP (most important) + NADH (More later) + glucose (sugar) ADP+ Pi  ATP with the human body turning over its own weight in ATP each day You get virtually all of the energy and the stuff which makes you, from the food that you eat.(You get a tiny bit from sunlight.) ATP http://tuberose.com/Dighttp://tuberose.com/Digestion.html

27. Need to know Chemical Bonding 4 types: what are they? – 100kT. Sharing of electrons. C-H 1. Covalent Is light enough to break covalent bond? 1um=1eV; kT=1/20eV. 1um= 20kT: close (yup) – varies tremendously, 100kT to few kT. + and – attract, but depends on solvent. Na+ Cl- = few kT (break up easily) 2. Ionic 3. Hydrogen – few kT, up to 5kT • Hydrogen attached to a very electro-negative elements, (O, N) causing the hydrogen to acquire a significant amount of positive charge. • Lone pair– electrons in relatively small space, very negative. • Result is H is (+) and O is (-). Will bind to other molecules –kT (weakest, but many of them together--significant). Two neutral atoms have instantaneous dipoles, and attract. 4. Van der Waals Neon: -246°C; Xenon: -108°C www.chemguide.co.uk/atoms/bonding/hbond.html#top

28. Fig. **. The rate of growth of single molecules has doubled every 2.2 years. In singulo Biophysics Single molecules, single cells, single species, single planets… Heterogeneity is the norm Men vs. women: height, sex organs Important to understand (prostate cancer, ovarian cancer It is only in last 10 (20) years that single molecule detection has been possible!

29. RNA vs. DNALife probably began as an RNA world • RNA more flexible than DNA (lack of an OH-bond in RNA!), can do catalysis as well as store information. • RNA not as stable as DNA, therefore not as good at genetic/ information storage. • Life probably began based on RNA.

30. Simple Lipids triglycerides http://lipidlibrary.aocs.org/Lipids/whatlip/index.htm

31. Carbohydrates Carbohydrates is the primary source of energy. Carbohydrates, which as the name implies, contains carbon and “hydrates,” that is, water. It’s formula is (CH2O)n, where n = 3-8 (Fig. **). As you can see from Fig. **, a carbohydrate also contain two or more –OH groups, and contains a C=O (known as an aldehyde or a ketone) The simplest case, where the carbohydrate cannot be broken down into simpler carbohydrates, is known as monosaccharides (from the Greek “mono” = one; “sacchar” meaning sugar). A polysaccharide consists of many monosaccharides. Carbohydrates are the most common source of energy for the human body. Protein and fats build tissue and cells, although people can live healthy lives without carbohydrates. This is because (excess) proteins and fats can be turned into an energy-source. Nevertheless, carbohydrates are the most straightforward way to get energy. People in the U.S. typically get 40% to 60% of their energy from carbohydrates, although 55% to 75% is considered more ideal. (http://simple.wikipedia.org/wiki/Carbohydrate)

32. Proteins

33. DNA, Carbohydrates. Lipid have isomersi.e. can have identical mass, but different spatial Arrangements. Note that virtually all have isomers. For example, in DNA can be made of A- or B- or some more bizarre types of structures; Carbohydrates, shown in (D), can have the exact same chemical make-up, but the order can be arranged, leading to different branches.

34. Chemical Bonds. All chemistry can be boiled down to plus and minus attracts, and plus and plus or negative and negative repel. You get negative charge from having more electrons than protons, and positive charges from having more protons than electrons. Within this general truism, chemists define 4 different types of chemical bonds. They are covalent bonds, ionic bonds, hydrogen bonds, and Van der Waal bonds. Covalent bonds are ones in which two atoms share an electron(s)

35. Proteins have a fair amount of complexity, but not too much. • Proteins are made of 20 monomers, called amino acids, which are strung together in a linear arrangement to form a biopolymer. Amino acids can be thought of as letters, and a protein as the word. Each amino acid varies only in their side-group; the straight-on part which is involved in connecting them to another amino acid on either end is the same. The biopolymer folds up in its native three dimensional structure and performs its function, whether that be mechanical support, being a truck to transport things, or being a catalyst to make reactions happen on a reasonable time-scale. Frauenfelder wanted to know whether there were any principles behind the mechanism of its folding. **But is this new physics??** **What about non-poissonian behaviour in cell growth?/** In a famous paper, where he studied the binding of carbon monoxide to myoglobin, he discovered the protein-folding landscape {Austin, 1973 #3357; Alberding, 1976 #3355}. While undoubtedly famous, it has been a mixed blessing. It has led people on the complex path to actually calculate how proteins folds, for which there has been consider progress. On the other hand, it has been a process which has taken four decades, with a long way to go!

36. Biological physics : under the mantle of complex physics, where the complexity comes from biology. But it can’t be too complicated, otherwise you never get anywhere. At present, to deal with biological systems in their living environment is much too complex. Eventually, it might not be so, but for now it is. One system which is amenable to quantitative reasoning, is the study of protein folding, a field Hans Frauenfelder , former Professor of Physics and founder of biological physics: became famous for. • Here proteins have a fair amount of complexity, but not too much. Proteins are made of 20 monomers, called amino acids, which are strung together in a linear arrangement to form a biopolymer. Amino acids can be thought of as letters, and a protein as the word. Each amino acid varies only in their side-group; the straight-on part which is involved in connecting them to another amino acid on either end is the same. The biopolymer folds up in its native three dimensional structure and performs its function, whether that be mechanical support, being a truck to transport things, or being a catalyst to make reactions happen on a reasonable time-scale. Frauenfelder wanted to know whether there were any principles behind the mechanism of its folding. **But is this new physics??** **What about non-poissonian behaviour in cell growth?/** In a famous paper, where he studied the binding of carbon monoxide to myoglobin, he discovered the protein-folding landscape {Austin, 1973 #3357; Alberding, 1976 #3355}. While undoubtedly famous, it has been a mixed blessing. It has led people on the complex path to actually calculate how proteins folds, for which there has been consider progress. On the other hand, it has been a process which has taken four decades, with a long way to go!

38. Organism have developed an amazing ability, fine tuned over a billion or-so of years of evolution, to make macromolecules. In fact, there are just a few types of macromolecules and each type of macromolecule is made from only a few types of monomers. This makes it fairly easy to learn them. But the length of a polymer can vary enormously--from a single one to 10 million or so, and hence the number of possible combinations that make up the polymer, is enormous. What adds to the complexity is that even if two polymers are made up of the exactly the same type of monomers and have the same order of monomers, there can be an enormous variation in their physical placement, known as isomers. Hence, from this simple alphabet, comes an incredibly complex story that is life. For example, imagine if an organism consists of just 4 different letters, or monomers. From this it can make a polymer—the word which is formed from the monomer alphabet. A polymer of N length can have 4N different polymers. In biology, the polymer can be a million monomers long, but let’s just say N equals 15--a very short polymer. So for this short polymer, and very simple language (English has 26 letters) that’s 415 = 230 = 1,073,741,824 or over a billion different combinations. And the range is actually much bigger because there are isomers of these polymers. An isomer is where you have the same number of atoms in the two polymers, but the arrangements of these atoms are different. For example, each monomer can be connected to another monomer either straight or bent fashion. Then there is 2 n4n = 8n = 23n= 245 =35,184,372,088,832 or over 35 trillion different polymers of length N=15. So, from a relatively simple set, one can get tremendous diversity.

39. What’s a (Bacterial) Cell made of ? It is mostly water and macromolecules Water is absorbed without any processing (as is inorganic ions and certain simple carbohydrates).

40. Structure of DNA A-T, G-C bond, line up Two strands are oppositely oriented Form a minor and major groove U RNA U 6 Å 12 Å

41. Structure of DNA A-T, G-C bond, line up Two strands are oppositely oriented Form a minor and major groove U RNA U

42. DNA: A-T 2 H bonds; G-C 3 H bonds Thymine Cytosine Hydrogen Bonds (2kT) Adenine Guanine G-Cmore stable than A-T Minor grove Major grove • – p stacking keeps it together (Grease); Phosphate negative charge makes it water soluble (Sort of like soap)

43. Biological Physics“Good” Problem: Protein Folding Unfolded  Folded Hans Frauenfelder , founder of biological physics. Inactive  Active Wolynes, PNAS, 1998 Main driving force : 1) Shield hydrophobic (black spheres) residues/a.a. from solvent/ water; 2) Formation of intramolecular hydrogen bonds. Probably forms a-helices and b-sheets (a lot of H-bonds) along pathway. Protein folding Landscape: Spend 4 centuries on it!