Transcript Slide 1

• An intriguing example of how chirally enriched amino
acids in the prebiotic world can generate sugars with Dconfiguration & with enantioenrichment:
Cordova et al. Chem. Commun., 2005, 2047-2049
The Model:
O
L-proline
OH
O
H
H
OBn
BnO
2-4 days
O
OH
+
OBn OBn
BnO
OBn
OBn
95-99% ee
>99% ee
hexose sugar
L-proline: a 2° amine; popular as an
organocatalyst because it forms enamines
readily
O
N
H
L-proline
OH
Mechanism: enamine formation
O
+
H
OBn
O
+
N
O
OH
H
N
H
OH
OBn
dilute
O
OH
+
N
O
OH
OBn
OBn
O
H
OBn
N
OH
OBn
CO2H participates as acid
OH
OBn
O
OBn
1st aldol product
(4C)
OH
O
O
OH
N
O
OH
+
OBn
OH
OBn
2nd proline-mediated
aldol reaction
OBn
BnO
O
OBn
OBn
OH
OH
BnO
BnO
OBn
OBn
benzyl protected allose
BnO
OBn
O
OBn
OH
Enantioenrichment
% ee of sugar vs % ee of AA
• Initially used 80% ee proline to
catalyze reaction → >99% ee
of allose
• Gradually decreased enatiopurity of proline
– Found that optical purity of
sugar did not decrease until
about 30% ee of proline!
– Non-linear relationship!
•  chiral amplification
– % ee out >> % ee in!
• Suggests that initial chiral pool was composed of amino
acids
• Chirality was then transferred with amplification to
sugars → “kinetic resolution”
• Could this mechanism have led to different sugars
diastereomers?
• Sugars →→ RNA world →→ selects for L-amino acids?
Ancient Amino Acids
(i.e., meteorites)
• Small peptides?
Ancient Peptides
Enzymes
Catalysis by Small Peptides
• Small peptides can also catalyze aldol reactions with
enantioenrichment (See Cordova et al. Chem. Commun. 2005,
4946)
H
O
O
O
OH
Catalytic Peptide
+
i.e.,
NO2
L-ala-L-ala
L-val-L-val
L-val-L-ala
NO2
81-96 % ee
• Found to catalyze formation of sugars
• It is clear that amino acids & small peptides are capable
of catalysis i.e., do not need a sophisticated protein!
From Amino Acids  Peptides
• Peptides are short oligomers of AAs (polypeptide ~ 2050 AAs); proteins are longer (50-3000 AAs)
O
+
H 3N
Ala
H2O
O
CH 3
+
H 3N
O
Cys
CH 3
O
O
N
H
+
+
H 3N
CH 2SH
O
petide bond
O
CH 2SH
H2O
• Reverse reaction is amide hydrolysis, catalyzed by
proteases
• At first sight, this is a simple carbonyl substitution
reaction, however, both starting materials & products are
stable:
– RCO2- -ve charge is stabilized by resonance
– Amides are also delocalized &  carbon & nitrogen are
sp2 (unlike an sp3 N in an amine):
O
.. C
O
N
N
C
H
sp2
C
C
H
• Primary structure: AA sequence with peptide bonds
• Secondary structure: local folding (i.e. -sheet & -helix)
-sheet
 helix
Amide bond: Formation & Degradation
O
+
R
O
H
OH
N R'
R
H
N
H
R'
+
H2O
• Thermodynamics
Overall rxn is ~ thermoneutral (Δ G ~ 0)
Removal of H2O can drive reaction to amide formation
In aqueous solution, reaction favors acid
• Kinetics
O
Very slow reaction
Forward:
+
R
O
Resonance stablilized
anion -stable & not
prone to nucleophilic
attack
H
+
H
N R'
X
H
Protonated--not a
nucleophile
O
Reverse:
R
N
H
R'
+
resonance stabilized:
most stable C=O
derivative
H2O
X
weak nucleophile
T.I = tetrahedral intermediate
O
Reaction Coordinate Diagram:
Large EA
for
forward
reaction
TS2
TS1
ΔG
R
OH
NH2+
Charge
separation
EA
T.I
EA
No resonance
Large EA
for
reverse
reaction
 HIGH
ENERGY!
How do we overcome the barrier?
1) Heat
NH4+
+
H2N
-N C O
O
H2N
First “biomimetic” synthesis
Disproved Vital force theory
But, cells operate at a fixed temperature!
2)
Activate the acid:
Activated acid
acid
+
H2O
• Activation of carboxylic acid
e.g.
O
PCl5
acid chloride
R
Cl
O
R
OH
P2O5
-H2O
O
O
anhydride
R
O
R
(Inorganic compound raises energy of acid)
Activation of carboxylic acid (towards nucleophilic attack) is one
of the most common methods to form an amide (peptide)
bond---in nature & in chemical synthesis!
• Why is the energy (of acid) raised?
• Recall carboxylic acid derivative reactivity:
O
R
O
>
Cl
R
O
O
O
>
R
R
SR'
O
>
R
O
O P
O
O
>
R
O
>
OR'
R
O
>
OH
NHR'
R
O
increasing stability
increasing reactivity
• Depends on leaving group:
O
– Inductive effects (EWG)
– Resonance in derivative
– Leaving group ability
Cl > O > S >
..
..
..
N
Cl
..
N > O > S > Cl
O
+
NHR
Cl- > -OCOR > -SR > -OR > -NHR
• Nature uses acyl phosphates, esters (ribosome) & thioesters
(NRPS)—more on this later
3)
Catalysis
•
•
•
Lowering of TS energy
Usually a Lewis acid
catalyst such as
B(OR)3
Another problem with AA’s
O
H2N
OH
HN
O
HO
O
NH
NH2
O
•
•
•
This doesn’t occur in nature
Easy to form 6 membered ring rather than peptide
Acid activation can give the same product
• With 20 amino acids  chaos!
• How do we control reaction to couple 2 AAs together
selectively & in the right sequence? & at room temp (in
vivo)?
• Biological systems & synthetic techniques employ
protection & activation strategies!
– For peptide bond formation
– Many different R groups on amino acids  potential for many
side reactions
HO
i.e.,
O
O
H2N
OH
H
N
H2N
OH
O
HO
SERINE
hydroxyl group is
a good nucleophile
& needs to be protected
BEFORE we make peptide
bond
OH
• Nature uses protection & activation as part of its strategy to
make proteins on the ribosome:
Nature uses an Ester to activate acid (protein synthesis):
Adenylation
O
O
H
N
H
O
Adenosine
O
O
O
P O P O P O
O
O
O
R
Activation
Formyl-AA
(methionine)
O
H
N
H
O
O
O P Adenosine
O
R
(raises energy of CO2H)
tRNA OH
Primary amine is protected
from further reaction
3'-OH terminus of
specific tRNA
sequence
O
H
N
H
O
O
tRNA
R
ester: more reactive
than an acid
H
N
H
O
O
O
H2N
O
R
O
tRNA
tRNA
H
R
O
H
N
O
R
R
O
N
H
O
AA 3
AA1
AA2 AA3 AA4...O
H2 O
polypeptide
Each AA is attached to its specific tRNA
tRNA
NH2
tRNA
• A specific example: tyrosyl-tRNA synthase (from tyr)
O
+
O
NH3
Adenosine
Good L.G. (PPi)
O
P O P O P O
O
O
O
O
O
+
O
O
NH3
O
anhydride-like
P
Adenosine
O
OH
OH
3 potential
nucleophiles!
3 potential
reactive P's
R
one of 20 AA's
tRNAtyr
O
B
only!
L-enantiomer only!
OH
R
O
+
B
3'-OH
only!
O
NH3
O
OH
OH
tRNA Tyr
OH
• Control!
– Only way to ensure specificity is to orient desired nucleophile
(i.e., CO2-) adjacent to desire electrophile (i.e., P)
What about Nonribosomal Peptide Synthase (NRPS)?
– Uses thioesters
Adenylation
O
O
H2N
H2N
O P Adenosine
O
R
O
R
O
HS
O
NH2
NRPS S
R
Activated thioester
NRPS
good
Lv group
O
potential Nu:
NH2
NRPS S
R
O
NRPS S
NH2
Activated thioester
O
H
N
NRPS S
R
NH2
O
hydrolysis
nonribosomal peptide
• Once again, we see selectivity in peptide bond formation
– As in the ribosome, the NRPS can orient the reacting centres in
close proximity to eachother, while physically blocking other sites
Chemical Synthesis of Peptides
• Synthesis of peptides is of great importance to chemistry
& biology
• Why synthesize peptides?
– Study biological functions (act as hormones, neurotransmitters,
antibiotics, anticancer agents, etc)
• Study potency, selectivity, stability, etc.
– Structural prediction
• Three-dimensional structure of peptides (use of NMR, etc.)
• How?
– Solution synthesis
– Solid Phase synthesis
– Both use same activation & protection strategy
e.g. isopenicillin N:
NH3+
-O2 C
• To study enzyme IPNS, we
need to synthesize
tripeptide (ACV)
• Small molecule → use
solution technique
• Synthesis (in soln) can be
long & low yielding
• But, can still produce
enough for study
O
SH
N
H
H
N
O
CO2-
L--aminoadipyl-L-cysteinyl-D-valine
(ACV)
isopenicillin N synthase
NH3+
O
H
N
-O2 C
S
N
O
Isopenicillin N
Plan for Synthesis:
NH3+
O
-O2 C
SH
N
H
O
H
N
CO2-
-aminoadipic acid
* NH +
3
-O2 C
*
O
*
N
Needs to be activated
cysteine
H
O
* Need protecting groups
*SH
H
N
CO2-
*
valine
Protection of Carboxylic acid:
OH
Ph
H2 N
CO2 H
H2 N
H+
O
O
Ph
heat
= OBn
(benzyl)
Val
Selective Protection of R group (thiol):
H2N
CO2H
BnCl
H2N
CO2H
NaOH
SH
Cys
S
• Both the amino group & carboxylate of cysteine need to
couple to another AA
– But, we can’t react all 3 peptides at once (must be stepwise)
–  we protect the amino group temporarily, then deprotect later
Protection of the Amine:
(BOC)2O = an anhydride
O
O
O
O
O
O
H2N
CO2H
SBn
O
H
N
CO2H
SBn
2X protection
=
BOC
H
N
CO2H
SBn
Now that we have our protected AA’s, we need to activate the
carboxylate towards coupling
O
O
H
N
H2N
CO2H
O
SBn
Ph
O
Activation & Coupling (see exp 6):
+
H
N C N
R
BOCHN
CO2-
DCC
Cy
SBn
O
N C N Cy
H
good Lv
group
DCC = dicyclohexylcarbodiimide = Coupling reagent that serves to
activate carboxylate towards nucleophilic attack