Welcome to 3FF3! Bio-organic Chemistry Jan. 7, 2008

Instructor: Adrienne Pedrech –ABB 417 – -Course website: Lectures: MW 8:30 F 10:30 (CNH/B107) –Office Hours: T 10:00-12:30 & F 1:00-2:30 or by appointment –Labs: 2:30-5:30 M ( ABB 302,306) **Note: course timetable says ABB217 2:30-5:30 F (ABB 306)  Every week except reading week (Feb ) & Good Friday (Mar. 21)  Labs start Jan. 7, 2008 (TODAY!)

For Monday 7 th & Friday 11 th Check-in, meet TA, safety and Lab 1 (Isolation of Caffeine from Tea) Lab manuals: Buy today! BEFORE the lab, read lab manual intro, safety and exp. 1 Also need: –Duplicate lab book (20B3 book is ok) –Goggles (mandatory) –Lab coats (recommended) –No shorts or sandals Obey safety rules; marks will be deducted for poor safety Work at own pace—some labs are 2 or 3 wk labs. In some cases more than 1 exp. can be worked in a lab period—your TA will provide instruction

Evaluation Assignments2 x 5% 10% Labs: - write up 15% - practical mark 5% Midterm 20% Final 50% Midterm test: Fri. Feb. 29, 2008 at 7:00 pm Make-up test: TBD Assignments: Feb.6 – Feb.13 Mar.7 – Mar.14 Note: academic dishonesty statement on outline-NO copying on assignments or labs ( exception when sharing results )

Texts: Dobson “Foundations of Chemical Biology,” (Optional- bookstore) Background & “Refreshers” An organic chemistry textbook (e.g. Solomons) A biochemistry textbook (e.g. Garrett) 2OA3/2OB3 old exam on web This course has selected examples from a variety of sources, including Dobson &: Buckberry “Essentials of Biological Chemistry” Dugas, H. "Bio-organic Chemistry" Waldman, H. & Janning, P. “Chemical Biology” Also see my notes on the website

What is bio-organic chemistry? Biological chem? Chemical bio? Chemical Biology: “Development & use of chemistry techniques for the study of biological phenomena” (Stuart Schreiber) Biological Chemistry: “Understanding how biological processes are controlled by underlying chemical principles” (Buckberry & Teasdale) Bio-organic Chemistry: “Application of the tools of chemistry to the understanding of biochemical processes” (Dugas) What’s the difference between these??? Deal with interface of biology & chemistry

BIOLOGYCHEMISTRY Simple organics eg HCN, H 2 C=O (mono-functional) Cf 20A3/B3 Biologically relevant organics: polyfunctional Life large macromolecules; cells—contain ~ 100, 000 different compounds interacting 1 ° Metabolism – present in all cell (focus of 3FF3) 2 ° Metabolism – specific species, eg. Caffeine (focus of 4DD3) CHEMISTRY: Round-bottom flask BIOLOGY: cell How different are they?

Exchange of ideas: Biology Chemistry Chemistry explains events of biology: mechanisms, rationalization Biology –Provides challenges to chemistry: synthesis, structure determination –Inspires chemists: biomimetics → improved chemistry by understanding of biology (e.g. enzymes)

Key Processes of 1° Metabolism Bases + sugars → nucleosides nucleic acids Sugars (monosaccharides) polysaccharides Amino acids proteins Polymerization reactions; cell also needs the reverse process We will look at each of these 3 parts: 1)How do chemists synthesize these structures? 2)How are they made in vivo? 3)Improved chemistry through understanding the biology: biomimetic synthesis

Properties of Biological Molecules that Inspire Chemists 1)Large → challenges: for synthesis for structural prediction (e.g. protein folding) 2)Size → multiple FG’s (active site) ALIGNED to achieve a goal (e.g. enzyme active site, bases in NAs) 3)Multiple non-covalent weak interactions → sum to strong, stable binding non-covalent complexes (e.g. substrate, inhibitor, DNA) 4)Specificity → specific interactions between 2 molecules in an ensemble within the cell

5) Regulated → switchable, allows control of cell → activation/inhibiton 6) Catalysis → groups work in concert 7) Replication → turnover e.g. an enzyme has many turnovers, nucleic acids replicates

Evolution of Life Life did not suddenly crop up in its element form of complex structures (DNA, proteins) in one sudden reaction from mono- functional simple molecules In this course, we will follow some of the ideas of how life may have evolved:

RNA World Catalysis by ribozymes occurred before protein catalysis Explains current central dogma: Which came first: nucleic acids or protein? RNA world hypothesis suggests RNA was first molecule to act as both template & catalyst: catalysis & replication

How did these reactions occur in the pre-RNA world? In the RNA world? & in modern organisms? CATALYSIS & SPECIFICITY How are these achieved? (Role of NON-COVALENT forces– BINDING) a) in chemical synthesis b) in vivo – how is the cell CONTROLLED? c) in chemical models – can we design better chemistry through understanding biochemical mechanisms?

Relevance of Labs to the Course Labs illustrate: 1)Biologically relevant small molecules (e.g. caffeine – Exp 1) 2)Structural principles & characterization (e.g. anomers of glucose, anomeric effect, diastereomers, NMR, Exp 2) 3)Cofactor chemistry – pyridinium ions (e.g. NADH, Exp 3 & 4) 4)Biomimetic chemistry (e.g. simplified model of NADH, Exp 3) 5)Chemical mechanisms relevant to catalysis (e.g. NADH, Exp 3)

6)Application of biology to stereoselective chemical synthesis (e.g. yeast, Exp 4) 7)Synthesis of small molecules (e.g. drugs, dilantin, tylenol, Exp 5,7) 8)Chemical catalysis (e.g. protection & activation strategies relevant to peptide synthesis in vivo and in vitro, Exp 6) All of these demonstrate inter-disciplinary area between chemistry & biology

Two Views of DNA 1)Biochemist’s view: shows overall shape, ignores atoms & bonds 2)chemist’s view: atom-by-atom structure, functional groups; illustrates concepts from 2OA3/2OB3

Biochemist’s View of the DNA Double Helix Major groove Minor groove

Chemist’s View

BASES Aromatic structures: –all sp 2 hybridized atoms (6 p orbitals, 6 π e - ) – planar (like benzene) N has lone pair in both pyridine & pyrrole  basic (H + acceptor or e - donor)

6 π electrons, stable cation  weaker acid, higher pKa (~ 5) & strong conj. base sp 3 hybridized N, NOT aromatic  strong acid, low pKa (~ -4) & weak conj. base Pyrrole uses lone pair in aromatic sextet → protonation means loss of aromaticity (BAD!) Pyridine’s N has free lone pair to accept H+  pyridine is often used as a base in organic chemistry, since it is soluble in many common organic solvents

The lone pair also makes pyridine a H-bond acceptor e.g. benzene is insoluble in H 2 O but pyridine is soluble: This is a NON-specific interaction, i.e., any H-bond donor will suffice

Contrast with Nucleic Acid Bases (A, T, C, G, U) – Specific! Evidence for specificity? Why are these interactions specific? e.g. G-C & A-T

Evidence? –If mix G & C together → exothermic reaction occurs; change in 1 H chemical shift in NMR; other changes  reaction occurring –Also occurs with A & T –Other combinations → no change! e.g. Guanine-Cytosine: Why? –In G-C duplex, 3 complementary H-bonds can form: donors & acceptors = molecular recognition

Can use NMR to do a titration curve: Favorable reaction because ΔH for complex formation = -3 x H-bond energy ΔS is unfavorable → complex is organized  3 H-bonds overcome the entropy of complex formation **Note: In synthetic DNAs other interactions can occur

Molecular recognition not limited to natural bases:  Create new architecture by thinking about biology i.e., biologically inspired chemistry! Forms supramolecular structure: 6 molecules in a ring

Synthesis of Bases (Nucleic) Thousands of methods in heterocyclic chemistry– we’ll do 1 example: –May be the first step in the origin of life… –Interesting because H-CN/CN - is probably the simplest molecule that can be both a nucleophile & electrophile, and also form C-C bonds

Mechanism?

Other Bases? ** Try these mechanisms!

Properties of Pyridine We’ve seen it as an acid & an H-bond acceptor Lone pair can act as a nucleophile:

Balance between aromaticity & charged vs non-aromatic & neutral!  can undergo REDOX reaction reversibly:

Interestingly, nicotinamide may have been present in the pre-biotic world: NAD or related structure may have controlled redox chemistry long before enzymes involved! electical discharge CH 4 + N 2 + H 2

Another example of N-Alkylation of Pyridines This is an S N 2 reaction with stereospecificity

References Solomons Amines: basicity ch.20 –Pyridine & pyrrole pp –NAD + /NADH pp 645-6, 537-8, Bases in nucleic acids ch. 25 Also see Dobson, ch.9 Topics in Current Chemistry, v 259, p 29-68

Sugar Chemistry & Glycobiology In Solomons, ch.22 (pp , ) Sugars are poly-hydroxy aldehydes or ketones Examples of simple sugars that may have existed in the pre-biotic world:

Most sugars, i.e., glyceraldehyde are chiral: sp 3 hybridized C with 4 different substituents The last structure is the Fischer projection: 1)CHO at the top 2)Carbon chain runs downward 3)Bonds that are vertical point down from chiral centre 4)Bonds that are horizontal point up 5)H is not shown: line to LHS is not a methyl group

In (R) glyceraldehyde, H is to the left, OH to the right  D configuration; if OH is on the left, then it is L D/L does NOT correlate with R/S Most naturally occurring sugars are D, e.g. D-glucose (R)-glyceraldehyde is optically active: rotates plane polarized light (def. of chirality) (R)-D-glyceraldehyde rotates clockwise,  it is the (+) enantiomer, and also d-, dextro-rotatory (rotates to the right- dexter)  (R)-D-(+)-d-glyceraldehyde & its enantiomer is: (S)-L-(-)-l-glyderaldehyde (+)/d & (-)/l do NOT correlate

Glyceraldehyde is an aldo-triose (3 carbons) Tetroses → 4 C’s – have 2 chiral centres  4 stereoisomers: D/L erythrose – pair of enantiomers D/L threose - pair of enantiomers Erythrose & threose are diastereomers: stereoisomers that are NOT enantiomers D-threose & D-erythrose: D refers to the chiral centre furthest down the chain (penultimate carbon) Both are (-) even though glyceraldehyde is (+) → they differ in stereochemistry at top chiral centre Pentoses – D-ribose in DNA Hexoses – D-glucose (most common sugar)

Reactions of Sugars 1)The aldehyde group: a)Aldehydes can be oxidized “reducing sugars” – those that have a free aldehyde (most aldehydes) give a positive Tollen’s test (silver mirror) b)Aldehydes can be reduced

c)Reaction with a Nucleophile Combination of these ideas  Killiani-Fischer synthesis: used by Fischer to correate D/L- glyceraldehyde with threose/erythrose configurations:

Reactions (of aldehydes) with Internal Nucleophiles Glucose forms 6-membered ring b/c all substituents are equatorial, thus avoiding 1,3-diaxial interactions

Can also get furanoses, e.g., ribose: Ribose prefers 5-membered ring (as opposed to 6) otherwise there would be an axial OH in the 6-membered ring

Why do we get cyclic acetals of sugars? (Glucose in open form is << 1%) a)Rearrangement reaction: we exchange a C=O bond for a stronger C-O σ bond  ΔH is favored b)There is little ring strain in 5- or 6- membered rings c)ΔS: there is some loss of rotational entropy in making a ring, but less than in an intermolecular reaction:1 in, 1 out. ** significant –ve ΔS!  ΔG = ΔH - TΔS Favored for hemiacetal Not too bad for cyclic acetal

Anomers Generate a new chiral centre during hemiacetal formation (see overhead) These are called ANOMERS –β-OH up –α-OH down –Stereoisomers at C1  diastereomers α- and β- anomers of glucose can be crystallized in both pure forms In solution, MUTAROTATION occurs

Mutatrotation

In solution, with acid present (catalytic), get MUTAROTATION: not via the aldehyde, but oxonium ion At equilibrium, ~ 38:62 α:β despite α having an AXIAL OH…WHY? ANOMERIC EFFECT

O lone pair is antiperiplanar to C-O σ bond  GOOD orbital overlap (not the case with the β- anomer) oxonium ion Anomeric Effect

Projections

More Reactions of Sugars 1)Reactions of OH group(s): a)Esterification: b)Ethers:

b) Ethers (con’t) c)Acetals

c) Acetals (con’t)

These reactions are used for selective protection of one alcohol & activation of another (protecting group chemistry) 1° alcohol is most reactive  protect first AZT

e.g, synthesis of sucrose (Lemieux, Alberta) Can only couple one way—if we don’t protect, get all different coupling patterns Yet nature does this all of the time: enzymes hold molecules together in the correct orientation, BUT the mechanism still goes through an oxonium ion (more on this later)

Selectivity of Anomer Formation in Glycosides Oxonium ion can often be attacked from both Re & Si faces to give a mixture of anomers. How do we control this?

This reaction provides a clue to how an enzyme might stabilize an oxonium ion (see later)