Transcript Origins of Sugars in the Prebiotic World

Origins of Sugars in the Prebiotic World
• One theory: the formose reaction (discovered by
Butterow in 1861)
O
Mineral catalysis
H
H
mixture of sugars,
including a small
amount of ribose
eg Ca(OH)2
formaldehyde
H
Mechanism?
H
HOH
O
H
HO
H
O
OH
O
O
H
O
*
O
O
OH
paraformaldehyde
H
H2O
slow, very
unfavorable
O
OH
O
O
O
H
OH
H
O
O
O
OH
n
*
OH
Con’t
O
O
O
OH
depolymerise
H
OH
O
glycolaldehyde:
simplest sugar &
a catalyst for
further rxns
H
Ca(OH)2
O
HO
HO
H
OH
O
-O
H
glyceraldehyde
H
H
ene-diloate
(enol)
pentoses, hexoses
via
ene-diolate
OH
O
H
OH
-OH
H
OH
OH
O
-O
O
OH
OH
OH
dihydroxyacetone
via
ene-diolate
H
O
OH
HO
OH
erythrose/threose
H
O
OH
-OH
2x
OH
HO
OH
erythrose/threose
retro-aldol
O
H
glycolaldehyde:
cycle back for
catalysis
• Today, similar reactions are catalyzed by thiazolium, e.g.,
Vitamin B1 (TPP), another cofactor:
• Cf Exp. 7: Benzoin
(PP) HO
S
condensation
H
• e.g.
N+
Py
H
OH
+
H
O
O
HO
OP
glycolaldehyde
OH
O
G3P
OH
HO
OP
D-xylulose-5-P
Mechanism? Uses thiazolium
O
HO
R
-OH
S
S
H
-
H
N+
N+
Py
Py
OH
R
OH
S
carbanion: zwitterionic;
stablized by +/- charge
interaction
N+
:B
H
OH
Py
N+ acidifies H
O
OP
OH
R
S
OH
N+
thiazolium anion
catalyst
regenerated
R
Py
HO
OH
OP
S
N+
Py
HO
+
O
HO
OH
OP
xylulose-5-P
HO
H
R
S
N
Py
OH
OH
enamine
• We have seen how the intermediacy of the resonancestablized oxonium ion accounts for facile substitution at
the anomeric centre of a sugar
• What about nitrogen nucleophiles?
CO2H
Many examples:
CO2H
RO
O
+
CO2H
N
quinolinate
N
NADH
CO2H
OH
RO
O
RO
O
OH
+
OPP
OH
OH
OH
OH
R = H or P
NH3
RO
O
+
NH2
nucleosides
OH
OH
Could this process have occurred in the prebiotic world?
•
Reaction of an oxonium ion with a nitrogenous base:
NUCLEOSIDES!
HO
OH
O
Mn+
Mineral days?
OH
O
Hydrothermal
vents?
OH
NH
N
H
&/or
apatite
HO
(mineral
phosphate)
O
O
NH
HO
+
O
O
N
O
O
OH
HO
O
P
OH
O
O
OH
OH
OH
OH
Thymidine (a nuclesoside)
OH
Activated leaving group:
CATALYSIS
•
Nucleosides are quite stable:
1)
2)
Weaker anomeric effect: N< O < Cl (low electronegativity of N)
N lone pair in aromatic ring  hard to protonate
O
1)
O
NH
NH
HO
O
HO
O
N
+
OH
O
N
O
-
Charge
separation:unfavorable,
since -ve charge is on
N, a less
electronegative group
OH
OH
OH
Anomeric effect: Cl > O > N (remember the glycosyl chloride
prefers Cl axial
O
2)
O
NH
NH
HO
O
N
O
H+
+
HO
O
X
OH
O
N
H
OH
OH
lone pair part of
aromatic sextet
OH
aromaticity destroyed
(i.e., pyridine & pyrrole)
• These effects stabilize the nucleoside making its
formation possible in the pre-biotic soup
• Thermodynamics are reasonably balanced
• However, the reaction is reversible
– e.g. deamination of DNA occurs ~ 10,000x/day/cell in vivo
– Deamination is due to spontaneous hydrolysis & by damage of
DNA by environmental factors
– Principle of microscopic reversibility: spontaneous reaction
occurs via the oxonium ion
Ribonucleosides & Deoxyribonucleosides
Ribonucleosides
• Contain ribose & found in RNA:
Cytosine
Uracil
Adenine
Guanine
+
+
+
+
Ribose
Ribose
Ribose
Ribose




Cytidine (C)
Uridine (U)
Adenosine (A)
Guanosine (G)
Deoxyribonucleosides
• Contain 2-deoxyribose, found in DNA
Cytosine
Thymine
Adenine
Guanine
+
+
+
+
2-dR
2-dR
2-dR
2-dR




2’-deoxycytidine (dC)
2’-deoxythymidine (dT)
2’-deoxyadenosine (dA)
2’-deoxyguanosine (dG)
Ribonucleosides
O
NH2
N
N
HO
O
OH
O
N
HO
O
OH
OH
NH2
H
N
O
N
HO
OH
OH
uridine (U)
cytidine (C)
O
O
N
N
N
N
HO
OH
O
OH
N
N
N
NH2
OH
guanosine (G)
adenosine (A)
Deoxyribonucleosides
O
NH2
N
N
HO
O
N
O
OH
2'-deoxycytidine (dC)
HO
O
N
NH2
H
N
O
OH
2'-deoxythymidine (dT)
HO
O
N
O
N
N
N
OH
2'-deoxyadenosine (dA)
HO
O
N
N
N
OH
2'-deoxyguanosine (dG)
NH2
Important things to Note:
• Numbering system:
– The base is numbered first (1,2, etc), then the sugar (1’, 2’, etc)
• Thymine (5-methyl uracil) replaces uracil in DNA
• Confusing letter codes:
– A represents adenine, the base
– A also represents adenosine, the nucleoside
– A also represents deoxyadenosine (i.e., in DNA sequencing,
where “d” is often omitted)
– A can also represent alanine, the amino acid
• Nucleoside + phoshphate  nucleotide
• In the modern world, enzymes (kinases) attach
phosphate groups
O
HO
O
A
-O
P
OH
OH
P
O
O
O
HO
A
OH
P
O
O
O
O
OH
OH
Adenosine-5'monophosphate (AMP)
OH
Adenosine-5'diphosphate (ADP)
O
HO
A
O
O
OH
P
P
O
O
O
P
O
O
O
Adenosine-5'triphosphate (ATP)
Energy source for cell
Central to metabolism
In the pre-RNA world, how might this happen?
P
O
O
A
O
OH
OH
Observation:
O
NH
HO
5'
clay
O
N
4'
5' phosphate + 3' phosphate + higher phosphates
(30 % + 50%)
O
(apatite)
1'
NUCLEOTIDES!
2'
3'
OH
OH
• Surprisingly easy to attach phosphate without needing
an enzyme
– One hypothesis: cyclo-triphosphate (explains preference for
triphosphate
O
O
O
P
P
O
O
O
O
O
P
ATP
O
HO
O
T
Primary OH?
sterics?
OH
OH
release of some
ring strain in
cylcotriphosphate
drives reaction?
• If correct, this indicates a central role for triphosphates of
nucleosides (NTPs) in early evolution of RNA (i.e.,
development of the RNA world)
• NTPs central to modern cellular biology
Triphosphates
• Triphosphates are reactive
– Attack by a nucleophile at P, P or P gives a
good resonance stabilized leaving group (can
also assisted by metal cation)
• Other examples where phosphorylation is
essential include:
– Glucose metabolism
O
O
O
O
– Enzyme regulation: CarbohydrateO P OH +
HO P O P O P O
O A
metabolism,
Lipid metabolism, receptors
O
O
O
O
Mg2+
OH
OH
OH
ADP
• If the nucleophile is the 3’-OH group of another NTP,
then a nucleic acid is generated: polymer of nucleotides
– Oligomers (“oligos”)  short length (DNA/RNA polymers of long
length)
HO
P
P
P
O
O
O
P
O
O
P
O
O
P
O
PPPO O
B
O
+
O
Nuc
OH
OH
Mg2+
PPPO O
B1
O
O
OH
O P O
trinucleotide
O
O
B2
a dinucleotide-5'-PPP
"oligo" (polymer)
OH
OH
Note that nature faces some problems:
1) Nucleophilic attack required by 3’-OH, not 2’-OH
2) Specific attack on P required
3) In a mixture of NTPs, get non-specific sequence
4) Reaction rate is slow
• Nucleic acids contain a regular array of bases, spaced
evenly along a backbone of phosphates & sugars
• Even spacing allows self-recognition,
– i.e., RNA short stretches form in which bases complement
one another
– tRNA folds into a specific conformation (more about tRNA
later)
– DNA: strand I and its reverse complement form a regular
sequence with bases paired through H-bonds
tRNA
Copyright 2006, John Wiley & Sons Publishers, Inc.
DNA
Template-Directed Synthesis in the PreBiotic Soup
OH
O
HO
N
O
O
N
H2N
N
NH
O
N
OH O
N
O
O
P
O
O
O
O
OH
O
O
N
O
NH
H2N
O
N
N
N
O
OH
P
N
OH
OH
OH
• Template-directed synthesis in the pre-biotic world allows
AMPLIFICATION due to MOLECULAR RECOGNITION
& rate acceleration results: an entropic effect!
• Now, catalyzed by enzymes:
– DNA polymerase makes DNA copy of a DNA template (i.e.,
replication)
– RNA polymerase makes RNA copy of a DNA template
(transcription)
Mechanism of Chain Elongation reaction
catalyzed by RNA polymerase
RO
DNA template strand
DNA template strand
RO
B1
O
B1
O
PP
OH
OH
O
O
PPPO
P O
B2
O
OH
OH
B2
O
O
OH
H
H
Mechanism of Chain Elongation reaction
catalyzed by DNA polymerase
template strand
RO
RO
B1
O
B1
O
PP
OH
H
O
O
PPPO
OH
P O
B2
O
O
H
H
B2
O
OH
H
• Viruses contain
– Reverse transcriptase (RT): makes a DNA copy of RNA genome
• Template strand = RNA, Product = DNA
– RNA synthetase: makes an RNA copy of RNA
• Template strand = RNA, Product = RNA
RNA as a Catalyst = Ribozymes
• Tom Cech & Sid Altman- Nobel Prize (1989)
• Ribozymes that catalyze many reactions are being
discovered
– i.e., cleavage of RNA (this is the reverse of synthesis)
Yeast tRNA
3'
5'
O
O
B
C60
17
O
-O Pb
O H
O
O
3'
5'
P
O
B
O
2+
C60
17
HO Pb
U59
O
O
P
O
O
HO
O
2+
U59
• This reaction is specific:
– Pb2+ binds to U59/C60 (if these are mutated  no binding)
– Cleavage is specific  requires 2’-OH at B17
– One of few systems where x-ray structure is available revealing
potential mechanism
• Another example: Can RNA catalyze addition of a base
to a sugar? YES!
see (on website):
Lau, M; Cadieux, K; Unrau, P. J. Am. Chem. Soc., 126, 1568615693
Chemical synthesis  random sequences
of RNA
a) Attach sugar, lacking base, to 3’
end
b) Few molecules react with base
to make nucleotide at 3’ end
c) Sort out those with base at 3’ end
d) Amplify (PCR), enrich pool & cycle
many times
Gives pure catalytic RNA!