Chem 239Amino Acids, Peptides, and Proteins

Representations:Classification20 common amino acidsFisher projections and D-L notation

Chemical Properties of Amino Acids:Acid-base behaviorSynthesis

Reactions of Amino Acids:Laboratory chemistry Biochemical reactions

Peptides:Nature of peptidesAnalysis: Composition, end-group, sequenceSynthesis: protection, bond-formationSynthesis: Solution vs solid-phase

Proteins:Secondary structuresTertiary and quarternary structuresCoenzymes

ClassificationAmino Acids

ContainContain——NHNH22

and and ——COCO22

HH

Actually Actually ——NH NH 3+3+

and and ——COCO22

––

Classified as Classified as αα, , ββ, , γγ, , etcetc. amino acids . amino acids according the carbon that bears the according the carbon that bears the nitrogen.nitrogen.

++HH33 NNCHCH22 CHCH22 CHCH22 COCO22

––

CC CC

OO

OO––

HH

HH

HH33 NN++

Naming Amino Acids

NHNH33

++

COCO22––

11--Aminocyclopropanecarboxylic acid:Aminocyclopropanecarboxylic acid:an an --amino acid that is anamino acid that is an intermediate in the biosynthesisintermediate in the biosynthesis of ethyleneof ethylene

++HH33 NNCHCH22 CHCH22 COCO22

––33--Aminopropanoic acid:Aminopropanoic acid:a a --amino acid that is one ofamino acid that is one of the structural units present inthe structural units present in coenzyme Acoenzyme A

++HH33 NNCHCH22 CHCH22 CHCH22 COCO22

–– 44--Aminobutanoic acidAminobutanoic acid::a a --amino acid involved amino acid involved in the transmission of in the transmission of nerve impulsesnerve impulses

The 20 Key Amino AcidsThe 20 Key Amino Acids

•

More than 700 amino acids in nature.

•

20 amino acids are the building blocks of proteins. All are α-amino acids.

•

Differ in the group attached to the α

carbon.

CC CC

OO

OO––

RRR

HH

HH33 NN++

Vary in:Vary in:

Size and shapeSize and shape

Electronic characteristicsElectronic characteristics

nonpolar side chains

polar but nonionized side chains

acidic side chains

basic side chains

The only achiral aaThe only achiral aa

Disrupts local protein structureDisrupts local protein structure

CC CC

OO

OO––

HH

HH

HH33 NN++

GlycineGlycine

(Gly or G)(Gly or G)

Small achiral side chainSmall achiral side chain

Nonpolar side chainNonpolar side chain

CC CC

OO

OO––

CHCH33

HH

HH33 NN++

AlanineAlanine (Ala or A)(Ala or A)

CC CC

OO

OO––

CH(CHCH(CH33 ))22

HH

HH33 NN++

ValineValine (Val or V)(Val or V)

CC CC

OO

OO––

CHCH22 CH(CHCH(CH33 ))22

HH

HH33 NN++

LeucineLeucine (Leu or L)(Leu or L)

CC CC

OO

OO––

CHCH33 CHCHCHCH22 CHCH33

HH

HH33 NN++

IsoleucineIsoleucine (Ile or I)(Ile or I)

Alkyl groups as side chainsAlkyl groups as side chains

Nonpolar and hydrophobic Nonpolar and hydrophobic

Often found in core of soluble proteinsOften found in core of soluble proteins

Nonpolar side chainNonpolar side chain

CC CC

OO

OO––

CHCH33 SCHSCH22 CHCH22

HH

HH33 NN++

MethionineMethionine (Met or M)(Met or M)

ProlineProline

CC CC

OO

OO––

CHCH22

HH

HH22 NN++

HH22 CCCC HH22

(Pro or P)(Pro or P)

Only secondary amineOnly secondary amine

Disrupts local protein structuresDisrupts local protein structures

AUG coded AUG coded

““startstart””

amino acidamino acid

Nonpolar side chainNonpolar side chain

PhenylalaninePhenylalanineCC CC

OO

OO––

CHCH22

HH

HH33 NN++

(Phe or F)(Phe or F)

CC CC

OO

OO––

CHCH22

HH

HH33 NN++

HH

TryptophanTryptophan

(Trp or W)(Trp or W)NN

Contribute to AContribute to A280280

nmnm

(along with Try and Phe)(along with Try and Phe)

Polar (nonPolar (non--ionizable) side chainionizable) side chain

CC CC

OO

OO––

CHCH22 OHOH

HH

HH33 NN++

SerineSerine

(Ser or S)(Ser or S)

CC CC

OO

OO––

CHCH33 CHOHCHOH

HH

HH33 NN++

ThreonineThreonine

(Thr or T)(Thr or T)

Can form hydrogen bondsCan form hydrogen bonds

Can be phosphorylatedCan be phosphorylated

Can be glycosylatedCan be glycosylated

Polar (nonPolar (non--ionizable) side chainionizable) side chain

Can form disulfidesCan form disulfides

CC CC

OO

OO––

CHCH22 SHSH

HH

HH33 NN++

CysteineCysteine

(Cys or C)(Cys or C)

CC

CC

OO

OO––

CHCH22 SS --HH

HH33 NN++

CC

CC

OO

OO––

HH

HH33 NN++

SCHSCH22

CystineCystine

Polar (nonPolar (non--ionizable) side chainionizable) side chain

TyrosineTyrosine

CC CC

OO

OO––

CHCH22

HH

HH33 NN++

OHOH

(Tyr or Y)(Tyr or Y)

AsparagineAsparagine

CC CC

OO

OO––

HH

HH33 NN++

HH22 NCCHNCCH22

OO

(Asn or N)(Asn or N)

GlutamineGlutamine

CC CC

OO

OO––

HH

HH33 NN++

HH22 NCCHNCCH22 CHCH22

OO(Gln or Q)(Gln or Q)

All can Form H-bonds

Can be phosporylated

Can be glycosylated

Ionizable (acidic) side chainsIonizable (acidic) side chains

CC CC

OO

OO––

HH

HH33 NN++

OCCHOCCH22

OO

––

Aspartic AcidAspartic Acid

(Asp or D)(Asp or D)

CC CC

OO

OO––

HH

HH33 NN++

OCCHOCCH22 CHCH22

OO

––Glutamic AcidGlutamic Acid

(Glu or E)(Glu or E)

All can form H-bondsAll can form ionic bonds with (+) charged residues

Ionizable (Basic) side chainsIonizable (Basic) side chains

CC CC

OO

OO––

CHCH22 CHCH22 CHCH22 CHCH22 NHNH33

HH

HH33 NN++

LysineLysine

++

(Lys or K)(Lys or K)

CC CC

OO

OO––

CHCH22 CHCH22 CHCH22 NHCNHNHCNH22

HH

HH33 NN++

ArginineArginine

++NHNH22

(Arg or R)(Arg or R)

HistidineHistidine

CC CC

OO

OO––

HH

HH33 NN++

CHCH22

(His or H)(His or H)

NN NHNH

All can form H-bondsAll can form ionic bonds with (-) charged residues

Some acetylated and methylated

Varying polarity of side chainsVarying polarity of side chains

Configurations of α-Amino Acids

HH33 NN++

HH

RR

COCO22––

All common All common amino acids amino acids

(except glycine) (except glycine) have at least one have at least one

chiral centerchiral center

Properties of GlycineProperties of Glycine

High melting point:

When heated to 233°C,

it decomposes before it melts

Solubility:

Soluble in water;

Not soluble in nonpolar solvent

OO

OHOHHH22 NCHNCH22 CC••••

••••

••••

•••• ••••

––••••

OO

OOHH33 NNCHCH22 CC ••••

••••

•••• ••••++

more consistent more consistent with thiswith this

than thisthan this

a a zwitterionzwitterion or or dipolar iondipolar ion

AcidAcid--Base Properties Base Properties of Glycineof Glycine

––••••

OO

OOHH33 NCHNCH22 CC ••••

••••

•••• ••••++

––••••

OO

OOHH22 NCHNCH22 CC ••••

••••

•••• ••••

••••

OO

OHOHHH33 NCHNCH22 CC++

••••

••••

•••• ••••

ppKKaa = 2.34= 2.34

ppKKaa = 9.60= 9.60

The The pI pI of of glycine is 5.97.glycine is 5.97.

CC CC

OO

OO––

HH

HH

HH33 NN++

GlycineGlycineppKKa1a1 = = 2.342.34 ppKKa2a2 == 9.609.60 ppI I == 5.975.97

Aspartic acidAspartic acid

ppKKa1a1 = = 1.881.88 ppKKa2a2 == 3.653.65 ppKKa3a3 == 9.609.60 ppI I == 2.772.77

HH33 NN CC CC

OO

OO––

HH++

OCCHOCCH22

OO

––

LysineLysine ppKKa1a1 = = 2.182.18 ppKKa2a2 == 8.958.95 ppKKa3a3 == 10.5310.53 ppI I == 9.749.74HH33 NN CC CC

OO

OO––

HH++

CHCH22 CHCH22 CHCH22 CHCH22 NHNH33

++

Synthesis of Amino AcidsSynthesis of Amino Acids

From -Halo Carboxylic Acids

CHCH33 CHCOHCHCOH

BrBr

OO

2NH2NH33++

HH22 OO

CHCH33 CHCOCHCO

NHNH33

OO

++

––

(65(65--70%)70%)

++ NHNH44 BrBr

Strecker Synthesis

NHNH44 ClCl

NaNaCNCN

CHCH33 CHCH

OO

CHCH33 CHCHCC

NHNH22

NN

CHCH33 CHCHCCOO

NHNH33

OO

++

–– (52(52--60%)60%)

1. H1. H22 O, HCl, heatO, HCl, heat

2. HO2. HO––

Using Diethyl Acetamidomalonate

1. NaOCH1. NaOCH22 CHCH33

2. 2. CC66 HH55 CHCH22 ClCl

OO OO

CHCH33 CHCH22 OCCCOCHOCCCOCH22 CHCH33

HHCHCH33 CCNHNH

OO

OO OO

CHCH33 CHCH22 OCCCOCHOCCCOCH22 CHCH33

CHCH22 CC66 HH55CHCH33 CCNHNH

OO

(90%)(90%)

HBr, HHBr, H22 O, heatO, heat

OO OO

HOCCCOHHOCCCOH

CHCH22 CC66 HH55HH33 NN++

OO OO

CHCH33 CHCH22 OCCCOCHOCCCOCH22 CHCH33

CHCH22 CC66 HH55CHCH33 CCNHNH

OO

OO

HCCOHHCCOH

CHCH22 CC66 HH55HH33 NN++(65%)(65%)

––COCO22

Reactions of Amino AcidsReactions of Amino Acids

Acylation of Amino GroupAcylation of Amino Group

OO

HH33 NNCHCH22 COCO––++++ CHCH33 COCCHCOCCH33

OO OO

CHCH33 CCNHNHCHCH22 COHCOH

OO OO

(89(89--92%)92%)

Fisher Esterification of Carboxyl GroupFisher Esterification of Carboxyl Group

++ CHCH33 CHCH22 OHOH

HClHCl

OO

HH33 NNCHCOCHCO––++

CHCH33

(90(90--95%)95%)

OO

HH33 NNCHCOCHCHCOCH22 CHCH33

++

CHCH33

––ClCl

Amino acids are detected by the formation of a purple color on treatment with ninhydrin.

OHOH

OO

OO

OHOH++

OO

HH33 NNCHCOCHCO––++

RR

OOOO

OO

NN

OO

––

OO

RCHRCH ++ COCO22 ++ HH22 OO ++

Biochemical Reactions of Amino AcidsBiochemical Reactions of Amino Acids

Transamination Transamination viavia

LL--Glutamic AcidGlutamic Acid

HOHO22 CCHCCH22 CHCH22 CHCOCHCO22––

NHNH33++

++ CHCH33 CCOCCO22 HH

OO

enzymesenzymes

HOHO22 CCHCCH22 CHCH22 CCOCCO22 HH

OO

++ CHCH33 CHCOCHCO22––

NHNH33++

Biosynthesis of LBiosynthesis of L--TyrosineTyrosine

CHCH22 CHCOCHCO22––

NHNH33++

OO22 , enzyme, enzyme

CHCH22 CHCOCHCO22––

NHNH33++

HHOO

DecarboxylationDecarboxylation

CHCH22 CHCHCOCO22––

NHNH33++

––COCO22 , enzymes, enzymes

CHCH22 CHCH2 2 NHNH22

NN

NN HH

NN

NN HH

PeptidesPeptides

CHCH33

OO

CC++

HH

CC OO––

HH33 NN

OO

CC

HH

HH

CCHH33 NN++

OO––

CHCH33

OO

CCHH33 NN++

HH

CC

OO

CCNN

HH

HH

CC OO––

HH

Two α-amino acids are joined by a peptide bond in alanylglycine.

It is a dipeptide.

AlanylglycineAlanylglycine

CHCH33

OO

CCHH33 NN++

HH

CC

OO

CCNN

HH

HH

CC OO––

HH

HH

OO

CCHH33 NN++

HH

CC

OO

CCNN

HH

CHCH33

CC OO––

HH

AlanylAlanylglycineglycineAlaAla——GlyGly

AAGG

GlycylGlycylalaninealanineGlyGly——AlaAla

GGAA

CHCH33

OO

CCHH33 NN++

HH

CC

OO

CCNN

HH

HH

CC OO––

HH

The peptide bond is characterized by a planar geometry.

Introduction to Peptide Structure DeterminationIntroduction to Peptide Structure Determination

Amino Acid Analysis

Acid-hydrolysis of the peptide (6 M HCl, 24 hr)

Separated by ion-exchange chromatography

Detected using ninhydrin.

Automated method; requires only 10-5

to 10-7 g of peptide.

End Group Analysis

Several chemical methods have been developed for identifying the N-terminus

FFOO22 NN

NONO22

NNHCHHCH22 CC NNHCHCOHCHCO

CHCH33

NNHCHCHCHC

CHCH22 CC66 HH55

HH22 NNCHCCHC

OO OOOOOO

CH(CHCH(CH33 ))22

––

++

Introduction to Peptide Structure DeterminationIntroduction to Peptide Structure Determination

Edman Degradation

1. Method for determining N-terminal amino acid.

2.

Can be done sequentially one residue at a time on the same sample. Usually one can determine the first 20 or so amino acids from the N-terminus by this method.

3. 10-10 g of sample is sufficient.

4. Has been automated.

The key reagent in the Edman degradation is phenyl isothiocyanate.

NN CC SS

Introduction to Peptide Structure DeterminationIntroduction to Peptide Structure Determination

peptidepeptideHH33 NNCHCCHC

OO

RR

++NNHHCC66 HH55 NN CC SS ++

CC66 HH55 NNHCHCNHNHCHCCHC

OO

RR

NNHH

SS

peptidepeptide

HClHCl

HH33 NN++

++CC66 HH55 NNHH CC

SSCC

NN CHCH

RR

OO

peptidepeptide

++ HH33 NN++

peptidepeptideCCCC

NN

HNHN CHCH

RR

OOSS

CC66 HH55

phenylthiohydantoin (PTH)phenylthiohydantoin (PTH)derivative.derivative.

phenyl isothiocyanate

Introduction to Peptide Structure DeterminationIntroduction to Peptide Structure Determination

Introduction to Peptide Structure DeterminationIntroduction to Peptide Structure Determination