Chemical Biology

A few important points

To understand chemical biology, you also need to know these subjects:organic chemistry, biochemistry, molecule biologyorganic chemistry, biochemistry, molecule biology

Learning method

Performance during the lectures (question & answer) will be incorporatedinto your final grade . Everybody is required to come to the class on time.Any excuse for absence has to be reported before the class.

Chapter 1 Peptide and ProteinChapter 1. Peptide and Protein

Outline

1. Chemical biology of polypeptidea. basic conceptsb. chemical synthesis of polypeptidec. peptide mimicsd. application of polypeptide and peptide drugs

2. Chemical synthesis of proteina. total synthesis of proteinb. semi-synthesis of protein

3. Protein modification and protein drugsa. protein modificationb protein drugsb. protein drugs

Basic Concepts

PolypeptideProtein Amino acid

amino acid-amino acidL-formR: side chain

Natural polypeptide and proteins consist by 20 kinds of L-form -amino acids

Chemical Biology of Polypeptide

Peptide and amino acid

polypeptideAmino acid

Polypeptide is the polymer of amino acids through amide bonds(peptide bonds)Polypeptide is the polymer of amino acids, through amide bonds(peptide bonds).Polypeptides usually contain less than 50 amino acids

Chemical Biology of Polypeptide

Classification of Peptide

By original: ribosomal peptide, non-ribosomal peptide, synthetic peptide

By backbone: linear peptide, cyclic peptide

By amino acid numbers: oligopeptide, polypeptide(>15 amino acid)

Chemical Biology of Peptide

Chemical synthesis of peptide:Chemical synthesis of peptide:

Amide bond formation

Chemical Biology of Peptide

Choosing a protecting groups for amino acids:

avoid of racemization solubility orthogonal

O

O

OHN

O

ONH

OHN

Boc Fmoc

tert-butoxycarbonyl 9-fluorenylmethoxycarbonyl

Solution-phase peptide synthesis

Chemical Biology of Polypeptide

F ti f tid b d f i idFormation of peptide bond from amino acids

selectiveselective protection activation

amide bond formation

selective deprotection

Chemical Biology of Peptide

S lid h tid th i1963 Merrifield: solid phase peptide synthesis (SPPS) 1984 Nobel Prize

Solid-phase peptide synthesis:

Xdeproection

Xresin

deproection

REPEATED

deproection

Chemical Synthesis of Protein

Steps for solid-phase peptide synthesis:1. Connect the C-terminal of N-protected amino acid to resin.2. Deprotection of N-protecting group.3. Amide bond formation with another N-protected amino acid.4. Repeat N-deprotection and amide coupling

Comparison of protecting groups in SPPS:

Type Resin Deprotectioncondition

Cleavage condition

Auto synthesis

strong acid

week acid

seldom

convenientWang Trt Rink HMPA

Merrifield, Pam, MBHA

Wang, Trt, Rink, HMPA

Chemical Synthesis of Protein

Cyclic peptide:

Ciclosporin

Peptide Mimics

Can polypeptide be used as drugs?

Easy to be hydrolyzed by enzymes so have lower stability, cant be taken orally.

seldom

Hydrophilic, make it hard to go through cell membrane.

Peptide Mimics: valuable for drug discovery and development

Peptoid

Peptide Peptoid

Peptide Mimics

-peptide :

Peptide Mimics

Application of polypeptide and peptide mimics :

Functional site of target proteing p Target identification and confirmation Ligand in high throughput screening Drug deliver system

Chemical Synthesis of Protein

Usually contain only natural amino acid, hard to introduce unnatural amino acid.

Main way to get a protein: biological expression

y y , Hard to control post-translational modification precisely Difficulties for biological expression of protein larger than 30kDa Difficult to express membrane protein or toxic proteins

Alternative: Chemical synthesis

Native chemical ligation (NCL) Native chemical ligation (NCL) Protein semisynthesis (eg. EPL)

Chemical Synthesis of Protein

Native Chemical Ligation :

1. Transthioesterification2. Thioester intermediate3. S N acyl shift

Catalyst if necessary

HSS

O

O ONa

2-mercaptoethanesulfonate

Chemical Synthesis of Protein

Semisynthesis of protein:

protein splicing

Chemical Synthesis of Protein

Semisynthesis of protein:

Thioester preparation by protein splicing p p y p p g

Chemical Synthesis of Protein

Express protein ligation (EPL)

Semisynthesis of protein:

p p g ( )

Chemical Synthesis of ProteinApplication of EPL:Application of EPL:

Synthesis of Ub-Histone H2B

Protein Modification and Protein Drugs

Protein Modification

Change the chemical structure of protein through chemical methods introduceChange the chemical structure of protein through chemical methods, introduce specific functional group and change its chemical function and biological properties, to study the relationship between protein structure and protein function and change the structure and function of protein with specificityfunction and change the structure and function of protein with specificity

High requirement for organic reaction

ReactivityChemo selectivityFunctional group compatibility

Protein Modification and Protein Drugs

Tyrosine modification

Protein Modification: Modification of protein residues

Tyrosine modification

good selectivityaromatic electrophilic substitution

OHprotein

aromatic electrophilic substitution (iodonation, nitration, )

J. Am. Chem. Soc., 2004, 126 (49), pp 1594215943

Protein Modification and Protein DrugsProtein Modification: Modification of protein residuesProtein Modification:

Lysine modification

Modification of protein residues

Protein Modification and Protein Drugs

Cysteine modification

Protein Modification:

t i

Modification of protein residues

Cysteine modification protein SH

1. C-S bond formation: alkylationy

R-X

2. S-S bond formation: reaction with disulfide

maleimidealkyl halide

2,2'-Dithiodipyridine

Protein Modification and Protein Drugs

N-terminal selective modification

Protein Modification: Modification of protein residues

C-terminal selective modification

Protein Modification and Protein Drugs

Modification of special sequence

Protein Modification: Modification of protein residues

Modification of special sequenceFLAsH

Protein Modification and Protein Drugs

Protein Modification:

Bio-orthogonal modifications

Ketone modification

g

Ketone modificationStaudinger ligationClick reaction

Protein Modification and Protein Drugs

Bio-orthogonal modifications

Staudinger azide-phosphine reaction

Protein Modification and Protein Drugs

Protein Modification: Bio-orthogonal modifications

Staudinger ligationStaudinger ligation traceless for chemoselective synthesis of amide bonds

Carolyn R. Bertozziy

Protein Modification and Protein Drugs

Protein Modification:

Bio-orthogonal modificationsgClick reaction

3+2 cycloaddition of azide and alkyney y

Protein Modification and Protein Drugs

Protein Modification:

Bio-orthogonal modificationsgClick reaction

Protein Modification and Protein Drugs

Protein Drugs:

Sources of protein drugs: IsolationDNA recombination and cell engineeringp g DNA recombination and cell engineering

Protein drugs as therapeutics:Protein drugs as therapeutics:

Peptide vaccine natural immune peptidep p synthetic peptide vaccine

Mono-cloned antibody (mAb)

Multifunctional protein drugs

Outline of Todays Lecture

1. Chemical biology of polypeptidebasic concepts (amino acid, peptide)chemical synthesis of polypeptide

(amide bond formation, protecting groups, SPPS)peptidomimetics (peptoid, -peptide)application of polypeptide and peptide drugs

2. Chemical synthesis of proteintotal synthesis of proteinsemi-synthesis of protein (NCL, EPL)

3. Protein modification and protein drugsprotein modification (side chain modification, bio-orthogonal modification)protein drugsprotein drugs

Homework

Get aware of natural amino acids:

1. Name and structure of natural amino acids.

2 Classification of the amino acids by side chain2. Classification of the amino acids by side chain.