Chapter 26: Amino Acids,Chapter 26: Amino Acids,
Peptides, Proteins, andPeptides, Proteins, and
Nucleic AcidsNucleic Acids...
Amino AcidsAmino Acids
In nature, the most common representativesIn nature, the most common representatives
are 2-amino ac...
Amino acids give rise to polyamides:Amino acids give rise to polyamides:
Polypeptides (proteins, enzymes)Polypeptides (pro...
• More than 500 natural amino acids knownMore than 500 natural amino acids known
• Among the 20 main amino acids, 8 cannot...
Used in reversible disulfide bridgingUsed in reversible disulfide bridging
MonosodiumMonosodium
glutamateglutamate
Amino acids are acidic and basic: Exist asAmino acids are acidic and basic: Exist as
zwitterionszwitterions
ppKKaa = 9-10=...
Structure depends on pH:Structure depends on pH:
PredominatesPredominates
at pH < 1at pH < 1
PredominatesPredominates
at p...
Isoelectric PointIsoelectric Point: When solution is charge: When solution is charge neutralneutral
[H[H33NCHCOOH] = [HNCH...
Synthesis of AminoSynthesis of Amino
AcidsAcids
1.1. Hell-Volhard-Zelinsky, then AminationHell-Volhard-Zelinsky, then Amin...
2. Gabriel Synthesis2. Gabriel Synthesis
Made from malonicMade from malonic
ester byester by
bromination, viabromination, ...
3. Strecker Synthesis3. Strecker Synthesis
Recall:Recall:
WhenWhen HCN is used in the presence of ammonia, e.g. NHHCN is u...
4. Synthesis of Enantiomerically Pure
Amino Acids
a. Reso-a. Reso-
utionution
b. Biological methodsb. Biological methods
RCCOOH
NH3, enzyme
C
H
COOHR
NH2
*
O
ReductiveReductive
aminationamination
Oxid...
Glutamic acid aminates otherGlutamic acid aminates other αα-oxoacids by redox exchange:-oxoacids by redox exchange:
RCCOOH...
c. Using Chiral Auxiliariesc. Using Chiral Auxiliaries
RCCOOH
O
NH3
Chiral,
enantiopure
hydride
C
H
COOHR
NH2
*
Or enantio...
PeptidesPeptides
Amino acids form peptide bonds
Dimer = dipeptide, trimer = tripeptide, and so on.Dimer = dipeptide, trime...
Rigidity and planarity of the peptide bondRigidity and planarity of the peptide bond
RigidRigid
spsp22
TransTrans
Amino endAmino end
to the leftto the left
Carboxy endCarboxy end
to the rightto the right
Main chainMain chain
RR,, R’R’,,...
Names:Names: Line up the amino acid names in sequence, fromLine up the amino acid names in sequence, from leftleft
to righ...
Found in all living cells, particularly in the lens of
the eye. Functions as a biological reducing agent
by enzymatic oxid...
Gramicidine S: Antibiotic (Eye Infection)Gramicidine S: Antibiotic (Eye Infection)
Two identical pentapeptides
joined head...
InsulinInsulin: For diabetes. Disulfide bonds: For diabetes. Disulfide bonds
The amino acid sequence is called theThe amino acid sequence is called the primaryprimary
structurestructure, the three di...
Sheets definedSheets defined
by shadedby shaded
areasareas
αα – Helix– Helix
Right-handed spiral held by intramolecular H bondsRight-handed spiral held by intramolecular H bonds
3.6...
Tertiary StructureTertiary Structure:: Further folding,
coiling, and aggregation. Denaturation is
the breakdown of this st...
Tertiary structure gives rise to pockets:Tertiary structure gives rise to pockets:
Active sites or binding sites that prov...
ProteinProtein
digestion =digestion =
AmideAmide
bondbond
hydrolysishydrolysis
FourFour
cooperativecooperative
effects toe...
Quaternary Structure:Quaternary Structure: Aggregation ofAggregation of
several unitsseveral units
ExampleExample::
Hemogl...
Protein (Polypeptide)Protein (Polypeptide)
Primary StructurePrimary Structure
DeterminationDetermination
Amino Acid Sequen...
ChromatographyChromatography
PaperPaper
AffinityAffinity ColumnColumn
Used after hydrolysis of polypeptides to componentUsed after hydrolysis of polypeptides to component
amino acidsamino acid...
3. Amino Acid Sequencing: One by one3. Amino Acid Sequencing: One by one
a.a. SangerSanger: Degrade from amino terminal en...
b.b. Edman degradationEdman degradation– leaves rest of– leaves rest of
chain intact, allows iterative procedurechain inta...
All phenylthiohydantoins of amino acids are known. EdmanAll phenylthiohydantoins of amino acids are known. Edman
degradati...
Enzymatic CleavageEnzymatic Cleavage
TrypsinTrypsin: Hydrolyzes only at carboxy end of: Hydrolyzes only at carboxy end of
...
Synthesis ofSynthesis of
PolypeptidesPolypeptides
Protecting groupsProtecting groups: Why? We need selectivity in: Why? We...
a. Amino end protection witha. Amino end protection with
PhenylmethoxycarbonylPhenylmethoxycarbonyl
(carbobenzoxy, Cbz)(ca...
b. Amino end protection withb. Amino end protection with
TertTert-butyloxycarbonyl-butyloxycarbonyl
(Boc)(Boc)
H3C C
O
O
H...
c. Carboxy Protection: Esterificationc. Carboxy Protection: Esterification
Simple methyl or ethyl esters. Alternatively, t...
Formation of the amide bond for peptides uses aFormation of the amide bond for peptides uses a
mild coupling reagent:mild ...
Mechanism:Mechanism:
1.1. ActivationActivation of carboxy group (recall activations toof carboxy group (recall activations...
2. Coupling with amino end of carboxy end protected2. Coupling with amino end of carboxy end protected
amino acid to give ...
AutomationAutomation: Merrifield solid-phase: Merrifield solid-phase
peptide synthesispeptide synthesis
Robert B. Merrifie...
Early mile stone:Early mile stone:
1966; Total1966; Total
synthesis ofsynthesis of
insulin with 51insulin with 51
amino ac...
DNA and RNA: NaturalDNA and RNA: Natural
Polymers Containing thePolymers Containing the
Blueprint of LifeBlueprint of Life...
Example: DNA chain.Example: DNA chain.
Backbone is a polymerBackbone is a polymer
of the sugar linked byof the sugar linke...
Bases: All aromatic heterocyclesBases: All aromatic heterocycles
ForFor
DNA:DNA:
For RNA: C, A, G andFor RNA: C, A, G and
...
NamingNaming the pieces:the pieces:
Sugar-base compound:Sugar-base compound:
AA nucleosidenucleoside
Sugar-base-phosphateS...
Primary structurePrimary structure
DNA forms extraordinarily long chains
(up to several centimeters) with
molecular weight...
Watson-Crick: Hydrogen bonding throughWatson-Crick: Hydrogen bonding through
complementary pairs: In DNA A-T, G-C (1:1 rat...
View down the helical axisView down the helical axis
Complementarity:Complementarity:
Replication of DNAReplication of DNA
In humans 2.9 billion base pairs; error rate 1 in 10 billion!In humans 2.9 billion ba...
Information storage: Peptides versusInformation storage: Peptides versus
nucleic acidsnucleic acids
Four letters: The four...
Base sequence contains information forBase sequence contains information for
protein synthesisprotein synthesis
DNADNA  ...
Three base code (Three base code (codoncodon): # of combinations): # of combinations
4433
= 64= 64  more than enough for...
Genetechnology: AGenetechnology: A
RevolutionRevolution
Deciphering theDeciphering the
sequence ofsequence of 2.92.9
billionbillion base pairsbase pairs
TheThe
HumanHuman
GenomeG...
The Future ?The Future ?
http://funnyjunk.com/pages/world.htm/http://funnyjunk.com/pages/world.htm/
Chapter26氨基酸肽蛋白质以及核酸
Chapter26氨基酸肽蛋白质以及核酸
Chapter26氨基酸肽蛋白质以及核酸
Chapter26氨基酸肽蛋白质以及核酸
Upcoming SlideShare
Loading in …5
×

Chapter26氨基酸肽蛋白质以及核酸

272 views

Published on

Published in: Technology, Business
0 Comments
0 Likes
Statistics
Notes
  • Be the first to comment

  • Be the first to like this

No Downloads
Views
Total views
272
On SlideShare
0
From Embeds
0
Number of Embeds
3
Actions
Shares
0
Downloads
9
Comments
0
Likes
0
Embeds 0
No embeds

No notes for slide
  • Affinity Chromatography
    The goal of affinity chromatography is to separate all the molecules of a particular specificity from the whole gamut of molecules in a mixture such as a blood serum. For example, the antibodies in a serum sample specific for a particular antigenic determinant can be isolated by the use of affinity chromatography.
    Step 1.
    An immunoadsorbent is prepared. This consists of a solid matrix to which the antigen (shown in blue) has been coupled (usually covalently). Agarose, sephadex, derivatives of cellulose, or other polymers can be used as the matrix.
    Step 2.
    The serum is passed over the immunoadsorbent. As long as the capacity of the column is not exceeded, those antibodies in the mixture specific for the antigen (shown in red) will bind (noncovalently) and be retained. Antibodies of other specificities (green) and other serum proteins (yellow) will pass through unimpeded.
    Step 3.
    Elution. A reagent is passed into the column to release the antibodies from the immunoadsorbent. Buffers containing a high concentration of salts and/or low pH are often used to disrupt the noncovalent interactions between antibodies and antigen. A denaturing agent, such as 8 M urea, will also break the interaction by altering the configuration of the antigen-binding site of the antibody molecule.
    Another, gentler, approach is to elute with a soluble form of the antigen. These compete with the immunoadsorbent for the antigen-binding sites of the antibodies and release the antibodies to the fluid phase.
    Step 4.
    Dialysis. The eluate is then dialyzed against, for example, buffered saline in order to remove the reagent used for elution.
    Link to another example of affinity chromatography.Link to a discussion of another chromatographic technique: exclusion chromatography.Welcome&amp;Next Search8 August 2003
  • Chapter26氨基酸肽蛋白质以及核酸

    1. 1. Chapter 26: Amino Acids,Chapter 26: Amino Acids, Peptides, Proteins, andPeptides, Proteins, and Nucleic AcidsNucleic Acids Aspartame, a dipeptideAspartame, a dipeptide Peptide linkage:Peptide linkage: An amideAn amide
    2. 2. Amino AcidsAmino Acids In nature, the most common representativesIn nature, the most common representatives are 2-amino acids (are 2-amino acids (αα-amino acids) with the-amino acids) with the general formulageneral formula RCH(NHRCH(NH22)COOH.)COOH. R = alkyl, acyl, amino, hydroxy, mercapto, sulfide,R = alkyl, acyl, amino, hydroxy, mercapto, sulfide, carboxy, guanidino, or imidazolyl groupscarboxy, guanidino, or imidazolyl groups H2N C COOH R H
    3. 3. Amino acids give rise to polyamides:Amino acids give rise to polyamides: Polypeptides (proteins, enzymes)Polypeptides (proteins, enzymes) Amide linkageAmide linkage For proteins,For proteins, nn ≥≥ 8000 and MW > 1,000,000. Proteins are8000 and MW > 1,000,000. Proteins are crucial for transport (Ocrucial for transport (O22, hemoglobin), energy storage,, hemoglobin), energy storage, catalysis, control of reactions, template for RNA/DNAcatalysis, control of reactions, template for RNA/DNA action, antibodies, etc.action, antibodies, etc. H2N O OH R N H H N R O R O n xx
    4. 4. • More than 500 natural amino acids knownMore than 500 natural amino acids known • Among the 20 main amino acids, 8 cannot beAmong the 20 main amino acids, 8 cannot be synthesized by the body (synthesized by the body (essential amino acidsessential amino acids)) Names:Names: We uWe use common names.se common names. αα-Stereocenter-Stereocenter usuallyusually SS (or, old nomenclature, L, from L-(or, old nomenclature, L, from L- glyceraldehyde)glyceraldehyde)
    5. 5. Used in reversible disulfide bridgingUsed in reversible disulfide bridging MonosodiumMonosodium glutamateglutamate
    6. 6. Amino acids are acidic and basic: Exist asAmino acids are acidic and basic: Exist as zwitterionszwitterions ppKKaa = 9-10= 9-10 ppKKaa = 2-3= 2-3 ZwitterionZwitterion Glycine as a zwitterionGlycine as a zwitterion HH33NCHNCH22COOCOO–– ++ H2N C H COOH R H2N C H COO RH - + :
    7. 7. Structure depends on pH:Structure depends on pH: PredominatesPredominates at pH < 1at pH < 1 PredominatesPredominates at pH ~ 6at pH ~ 6 PredominatesPredominates at pH > 13at pH > 13 Compare pCompare pKKaa CHCH33COOHCOOH == 4.744.74; the -NH; the -NH33 ++ groupgroup acidifiesacidifies
    8. 8. Isoelectric PointIsoelectric Point: When solution is charge: When solution is charge neutralneutral [H[H33NCHCOOH] = [HNCHCOOH] = [H22NCHCOO ]NCHCOO ] RR RR -- At this point pH =At this point pH = Cf. pCf. pKKaa CHCH33 NHNH33 ++ == 10.6210.62; -CO; -CO22 -- alsoalso acidifies, by inductionacidifies, by induction ++
    9. 9. Synthesis of AminoSynthesis of Amino AcidsAcids 1.1. Hell-Volhard-Zelinsky, then AminationHell-Volhard-Zelinsky, then Amination (racemic)(racemic) Yields not greatYields not great Better:Better: GabrielGabriel Synthesis ( RSynthesis ( RXX →→ RNHRNH22))
    10. 10. 2. Gabriel Synthesis2. Gabriel Synthesis Made from malonicMade from malonic ester byester by bromination, viabromination, via enolateenolate General:General:
    11. 11. 3. Strecker Synthesis3. Strecker Synthesis Recall:Recall: WhenWhen HCN is used in the presence of ammonia, e.g. NHHCN is used in the presence of ammonia, e.g. NH44CN orCN or NHNH44Cl/NaCN, we get the corresponding amino cyanide:Cl/NaCN, we get the corresponding amino cyanide: Adolf Strecker (1822–1871) StreckerStreckerSegoviaSegovia
    12. 12. 4. Synthesis of Enantiomerically Pure Amino Acids a. Reso-a. Reso- utionution
    13. 13. b. Biological methodsb. Biological methods RCCOOH NH3, enzyme C H COOHR NH2 * O ReductiveReductive aminationamination OxidationOxidation
    14. 14. Glutamic acid aminates otherGlutamic acid aminates other αα-oxoacids by redox exchange:-oxoacids by redox exchange: RCCOOH C H COOHR N * NH N HH R CONH2 : H+ HH Mechanism:Mechanism: Hydride transferHydride transfer from onlyfrom only one sideone side ofof double bond indouble bond in chiralchiral environmentenvironment of theof the enzymeenzyme
    15. 15. c. Using Chiral Auxiliariesc. Using Chiral Auxiliaries RCCOOH O NH3 Chiral, enantiopure hydride C H COOHR NH2 * Or enantio-Or enantio- selectiveselective StreckerStrecker synthesissynthesis
    16. 16. PeptidesPeptides Amino acids form peptide bonds Dimer = dipeptide, trimer = tripeptide, and so on.Dimer = dipeptide, trimer = tripeptide, and so on. The chain is arranged in space by H bonding,The chain is arranged in space by H bonding, electrostatic attractions, hydrophobic-electrostatic attractions, hydrophobic- hydrophilic interactions (with water), and rigidityhydrophilic interactions (with water), and rigidity of the amide bond.of the amide bond.
    17. 17. Rigidity and planarity of the peptide bondRigidity and planarity of the peptide bond RigidRigid spsp22 TransTrans
    18. 18. Amino endAmino end to the leftto the left Carboxy endCarboxy end to the rightto the right Main chainMain chain RR,, R’R’,, R”R”, etc. are called the, etc. are called the side chainsside chains.. All stereocenters are assumed to beAll stereocenters are assumed to be S.S.
    19. 19. Names:Names: Line up the amino acid names in sequence, fromLine up the amino acid names in sequence, from leftleft to right, each ending in “yl”, until reaching the terminus.to right, each ending in “yl”, until reaching the terminus. Examples of peptidesExamples of peptides Three letter codeThree letter code NutrasweetNutrasweet 200 times as sweet as table200 times as sweet as table sugar, only 4 cal/g vs 4 kcal/g.sugar, only 4 cal/g vs 4 kcal/g. Compare: Fat 9 kcal/g,Compare: Fat 9 kcal/g, chocolate 3 kcal/g.chocolate 3 kcal/g.
    20. 20. Found in all living cells, particularly in the lens of the eye. Functions as a biological reducing agent by enzymatic oxidation of the cysteine mercapto unit to the disulfide-bridged dimer. Not a typical peptide bond: Glutamic acid residue makes amide bond with the γ-carboxy group (γ-Glu). GlutathioneGlutathione
    21. 21. Gramicidine S: Antibiotic (Eye Infection)Gramicidine S: Antibiotic (Eye Infection) Two identical pentapeptides joined head to tail. !! Rare, lowerRare, lower homolog of lysinehomolog of lysine
    22. 22. InsulinInsulin: For diabetes. Disulfide bonds: For diabetes. Disulfide bonds
    23. 23. The amino acid sequence is called theThe amino acid sequence is called the primaryprimary structurestructure, the three dimensional arrangement is, the three dimensional arrangement is thethe secondary, tertarysecondary, tertary, and, and quaternaryquaternary structure.structure. Secondary Structure:Secondary Structure: H-BondingH-Bonding Pleated Sheet StructurePleated Sheet Structure
    24. 24. Sheets definedSheets defined by shadedby shaded areasareas
    25. 25. αα – Helix– Helix Right-handed spiral held by intramolecular H bondsRight-handed spiral held by intramolecular H bonds 3.6 Amino acids per turn; repeat distance 5.4 Å3.6 Amino acids per turn; repeat distance 5.4 Å
    26. 26. Tertiary StructureTertiary Structure:: Further folding, coiling, and aggregation. Denaturation is the breakdown of this structure. Example:Example: SuperhelixSuperhelix Typical of fibrous proteins, such as myosin (in muscle), fibrin (in blood clots), and α- keratin (in hair, nails, and wool).
    27. 27. Tertiary structure gives rise to pockets:Tertiary structure gives rise to pockets: Active sites or binding sites that provide perfect fit for substrates, e.g., drugs. Example:Example: Digestive enzyme chymotrypsinDigestive enzyme chymotrypsin
    28. 28. ProteinProtein digestion =digestion = AmideAmide bondbond hydrolysishydrolysis FourFour cooperativecooperative effects toeffects to facilitatefacilitate protonproton shuttleshuttle
    29. 29. Quaternary Structure:Quaternary Structure: Aggregation ofAggregation of several unitsseveral units ExampleExample:: HemoglobinHemoglobin
    30. 30. Protein (Polypeptide)Protein (Polypeptide) Primary StructurePrimary Structure DeterminationDetermination Amino Acid SequencingAmino Acid Sequencing 1.1. Break S-S bridges by oxidationBreak S-S bridges by oxidation  purify piecespurify pieces RR11 –S–S–S–R–S–R22 RR11 SOSO33H + RH + R22 SOSO33HH Purification is achieved byPurification is achieved by various forms ofvarious forms of chromatography: Dialysis,chromatography: Dialysis, gel-filtration, ion-exchange,gel-filtration, ion-exchange, electrophoresis, affinityelectrophoresis, affinity chromatography.chromatography.
    31. 31. ChromatographyChromatography PaperPaper AffinityAffinity ColumnColumn
    32. 32. Used after hydrolysis of polypeptides to componentUsed after hydrolysis of polypeptides to component amino acidsamino acids (6 N HCl, 110°C, 24 h). Column chromatography separates and detects them. 2. Amino Acid Analyzer2. Amino Acid Analyzer Various amino acidsVarious amino acids Integration gives relative amount of type of amino acid.Integration gives relative amount of type of amino acid. Now we know composition. What about the sequence?Now we know composition. What about the sequence?
    33. 33. 3. Amino Acid Sequencing: One by one3. Amino Acid Sequencing: One by one a.a. SangerSanger: Degrade from amino terminal end: Degrade from amino terminal end Analyze: All dinitrophenylAnalyze: All dinitrophenyl derivatives of amino acidsderivatives of amino acids are knownare known HCl,HCl, ΔΔ:: Hydrolyzes everythingHydrolyzes everything Nucleophilic substitutionNucleophilic substitution O2N NO2 F H2N H2 C C CH O R O + O2N NO2 N H O2N NO2 N H H2 C C OH O : H N H2 C C CH O R O H N
    34. 34. b.b. Edman degradationEdman degradation– leaves rest of– leaves rest of chain intact, allows iterative procedurechain intact, allows iterative procedure PhenylPhenyl isothiocyanate;isothiocyanate; like O=C=Olike O=C=O TransamidationTransamidation
    35. 35. All phenylthiohydantoins of amino acids are known. EdmanAll phenylthiohydantoins of amino acids are known. Edman degradation turns unreliable after ~50 amino acids (build up ofdegradation turns unreliable after ~50 amino acids (build up of impurities). Problem solved by chopping up large peptidesimpurities). Problem solved by chopping up large peptides selectively into smaller (< 50 amino acids) pieces usingselectively into smaller (< 50 amino acids) pieces using enzymesenzymes.. up to 50 cyclesup to 50 cycles
    36. 36. Enzymatic CleavageEnzymatic Cleavage TrypsinTrypsin: Hydrolyzes only at carboxy end of: Hydrolyzes only at carboxy end of Arg and Lys, e.g.:Arg and Lys, e.g.: ChymotrypsinChymotrypsin: Hydrolyzes only at carboxy: Hydrolyzes only at carboxy end of Phe, Trp, Tyr, e.g.:end of Phe, Trp, Tyr, e.g.: Trp-Glu-ArgTrp-Glu-Arg Phe-Phe-LysPhe-Phe-Lys Ala-ValAla-Val TrpTrp Glu-Arg-PheGlu-Arg-Phe PhePhe Lys-Ala-ValLys-Ala-Val ThermolysinThermolysin: Hydrolyzes amino end of: Hydrolyzes amino end of Leu, Ile, Val, e.g.:Leu, Ile, Val, e.g.: Trp-Glu-Arg-Phe-Phe-Lys-AlaTrp-Glu-Arg-Phe-Phe-Lys-Ala ValVal Followed by findingFollowed by finding overlapoverlap sequences to establish connectivity in originalsequences to establish connectivity in original
    37. 37. Synthesis ofSynthesis of PolypeptidesPolypeptides Protecting groupsProtecting groups: Why? We need selectivity in: Why? We need selectivity in building up the peptide sequence. Consider thebuilding up the peptide sequence. Consider the synthesis of glycylalanine by dehydration of thesynthesis of glycylalanine by dehydration of the component amino acids:component amino acids: Hence, amino end of Gly and carboxy end of Ala have to be protected.Hence, amino end of Gly and carboxy end of Ala have to be protected. H2N CH2 C OH O H2N CH C CH3 OH O +
    38. 38. a. Amino end protection witha. Amino end protection with PhenylmethoxycarbonylPhenylmethoxycarbonyl (carbobenzoxy, Cbz)(carbobenzoxy, Cbz) CH2 O O Like the hydrogenolysis of benzylic ethersLike the hydrogenolysis of benzylic ethers UnstableUnstable
    39. 39. b. Amino end protection withb. Amino end protection with TertTert-butyloxycarbonyl-butyloxycarbonyl (Boc)(Boc) H3C C O O H3C CH3 (via(via terttert-Bu cation)-Bu cation)
    40. 40. c. Carboxy Protection: Esterificationc. Carboxy Protection: Esterification Simple methyl or ethyl esters. Alternatively, to avoid base (or acid)Simple methyl or ethyl esters. Alternatively, to avoid base (or acid) hydrolysis,hydrolysis, benzyl esters.benzyl esters. H2N CH C CH2 OHO HN OH + H+ H2N CH C CH2 OO HN H2N CH C CH2 OO HN H2, Pd H2N CH C CH2 OHO HN CH3 + ProtectionProtection DeprotectionDeprotection TrpTrp Trp-OBzTrp-OBz
    41. 41. Formation of the amide bond for peptides uses aFormation of the amide bond for peptides uses a mild coupling reagent:mild coupling reagent: DicycloheylcarbodiimideDicycloheylcarbodiimide (DCC)(DCC) as a dehydrating speciesas a dehydrating species
    42. 42. Mechanism:Mechanism: 1.1. ActivationActivation of carboxy group (recall activations toof carboxy group (recall activations to alkanoyl halides or anhydrides) ofalkanoyl halides or anhydrides) of NN-protected amino acid-protected amino acid H N CH C CH3 OH O N C N + δ+ :: H3C C O OH3C CH3 HN CH C CH3 O O CH3 C OO H3C H3C NH C N Looks like an anhydrideLooks like an anhydride
    43. 43. 2. Coupling with amino end of carboxy end protected2. Coupling with amino end of carboxy end protected amino acid to give dipeptideamino acid to give dipeptide Peptide bondPeptide bond : HN CH C CH3 O O CH3 C OO H3C H3C NH C N H2N CH2 C OCH3 O HN CH C CH3 O CH3 C OO H3C H3C N H CH2 C OCH3O O NH C HN +
    44. 44. AutomationAutomation: Merrifield solid-phase: Merrifield solid-phase peptide synthesispeptide synthesis Robert B. Merrifield b. 1921 Advantages: Robotic “custom made” assembly ofAdvantages: Robotic “custom made” assembly of any polypeptide; products on the polymer areany polypeptide; products on the polymer are isolated and purified by simple filtration andisolated and purified by simple filtration and washing.washing.
    45. 45. Early mile stone:Early mile stone: 1966; Total1966; Total synthesis ofsynthesis of insulin with 51insulin with 51 amino acids;amino acids; 5000 operations5000 operations carried out in acarried out in a few days.few days. ModernModern extensions:extensions: CombinatorialCombinatorial chemistry andchemistry and total synthesistotal synthesis of complexof complex molecules onmolecules on solid supports.solid supports.
    46. 46. DNA and RNA: NaturalDNA and RNA: Natural Polymers Containing thePolymers Containing the Blueprint of LifeBlueprint of Life Life (in this context) is theLife (in this context) is the synthesis ofsynthesis of proteinsproteins, which run, which run our (any) body. The informationour (any) body. The information is stored in DNA:is stored in DNA: DeoxyriboseDeoxyribose nucleic acidnucleic acid The information is “read”The information is “read” (expressed) with the help of(expressed) with the help of RNA:RNA: Ribonucleic acidRibonucleic acid
    47. 47. Example: DNA chain.Example: DNA chain. Backbone is a polymerBackbone is a polymer of the sugar linked byof the sugar linked by phosphate groups as aphosphate groups as a diester.diester. ““Monomer”Monomer” is called ais called a nucleotidenucleotide Structure of Nucleic AcidsStructure of Nucleic Acids P OH HO OH O PhosphoricPhosphoric acidacid The informationThe information lies in thelies in the sequence of thesequence of the bases attachedbases attached to the anomericto the anomeric carbon:carbon: ThereThere are four bases.are four bases.
    48. 48. Bases: All aromatic heterocyclesBases: All aromatic heterocycles ForFor DNA:DNA: For RNA: C, A, G andFor RNA: C, A, G and Instead ofInstead of thyminethymine To visualize theTo visualize the aromaticity in the cyclicaromaticity in the cyclic amides, formulate theamides, formulate the dipolar resonance form,dipolar resonance form, e.g. cytosine:e.g. cytosine: N N H NH2 O N N NH2 O H
    49. 49. NamingNaming the pieces:the pieces: Sugar-base compound:Sugar-base compound: AA nucleosidenucleoside Sugar-base-phosphateSugar-base-phosphate monomer piece ofmonomer piece of polymer:polymer: AA nucleotidenucleotide
    50. 50. Primary structurePrimary structure DNA forms extraordinarily long chains (up to several centimeters) with molecular weights of as high as 150 billion. Like proteins, they adopt secondary and tertiary structures: Double helix! Human chromosomesHuman chromosomes
    51. 51. Watson-Crick: Hydrogen bonding throughWatson-Crick: Hydrogen bonding through complementary pairs: In DNA A-T, G-C (1:1 ratio)complementary pairs: In DNA A-T, G-C (1:1 ratio) AA TT CCGG Gives rise to double helix.......Gives rise to double helix....... 27.7 kcal mol-1 15.1 kcal mol-1
    52. 52. View down the helical axisView down the helical axis Complementarity:Complementarity:
    53. 53. Replication of DNAReplication of DNA In humans 2.9 billion base pairs; error rate 1 in 10 billion!In humans 2.9 billion base pairs; error rate 1 in 10 billion!
    54. 54. Information storage: Peptides versusInformation storage: Peptides versus nucleic acidsnucleic acids Four letters: The four bases R = 20 letters: The 20 natural amino acids
    55. 55. Base sequence contains information forBase sequence contains information for protein synthesisprotein synthesis DNADNA  mRNAmRNA  polypeptidepolypeptide Messenger RNAMessenger RNA mRNA copies a piece of DNA to be used to construct a particular peptide;mRNA copies a piece of DNA to be used to construct a particular peptide; transfer RNA (tRNA) delivers the amino acids, and a catalyst (ribosome)transfer RNA (tRNA) delivers the amino acids, and a catalyst (ribosome) puts the peptide together, following the blueprint: Three base sequencesputs the peptide together, following the blueprint: Three base sequences (codons) that translate into a specific amino acid.(codons) that translate into a specific amino acid.
    56. 56. Three base code (Three base code (codoncodon): # of combinations): # of combinations 4433 = 64= 64  more than enough for 20 amino acidsmore than enough for 20 amino acids
    57. 57. Genetechnology: AGenetechnology: A RevolutionRevolution
    58. 58. Deciphering theDeciphering the sequence ofsequence of 2.92.9 billionbillion base pairsbase pairs TheThe HumanHuman GenomeGenome
    59. 59. The Future ?The Future ? http://funnyjunk.com/pages/world.htm/http://funnyjunk.com/pages/world.htm/

    ×