Nucleic Acids in Chemistry and Biology

formation from phosphorus to the nucleophile and the leaving group. If this is greater in the transition state
than in the starting state (i.e.greater than one), then the transition state is considered associative (starting
to resemble the intermediate in Figure 3.44a); if it is less than one, then the transition state is dissociative
in character (starting to resemble the intermediate in Figure 3.44c). This issue is most important for
enzymes, which have evolved to stabilize transition states effectively, and considerable progress has been
made through the use of ‘metaphosphate’ mimics, such as AlF 3 and MgF 3 , in X-ray structures of
nucleotide complexes with enzymes that utilise ATP or GTP.

3.2.2.1 Hydrolysis of Alkyl Triesters. Trimethyl phosphate is hydrolysed in alkaline solution in an

SN2(P) process (kOH1.6 10 ^4 M^1 s^1 at 25°C). With H 218 O as solvent, no Me^18 OH is formed, which
shows that the reaction involves exclusively P O cleavage. Other ‘hard’ nucleophiles such as Freact
similarly, and indeed, fluoride catalysis of the trans-esterification of triesters is a useful process (Figure 3.45).
The intra-molecular migration of phosphorus in a triester to a vicinal hydroxyl group is especially easy
and must be avoided in the synthesis of oligoribonucleotide and inositol phosphate precursors.
Trimethyl phosphate is hydrolysed extremely slowly in neutral and acidic conditions (kw 2 10 ^8 s^1
at 25°C) with C O cleavage. Soft nucleophiles, such as RS, Bror I, also dealkylate phosphate triesters
with C O cleavage (Figure 3.46a). Such reactions are typical SN2 processes and show a clear preference for
dealkylation in the order of Me Et R 2 CH. This characteristic is particularly well exploited in the thio-
phenolate deprotection of methyl phosphate triesters used in the phosphodiester chemistry of oligonu-
cleotide synthesis (Section 4.1) (Figure 3.46b). However in this case, some (
20%) oligonucleotide chain
cleavage can result through attack of thiophenolate at the 5-carbon of the 3-nucleotide unit.
Alkyl phosphate triesters are sensitive to
-elimination processes as has been exploited for the selective
deprotection of phosphate triesters in oligonucleotide synthesis (Section 4.1). The 2-cyanoethyl group
possesses an acidic
-hydrogen atom and may be removed by
-elimination mechanism under mildly

Nucleosides and Nucleotides 103

O

(MeO) 2 P

O

(MeO) 2 P

OBn

O

(MeO) 2 P

F

O

HO (MeO) 3 P=O F BnOH

Figure 3.45 P O cleavage reactions with hard nucleophiles for triesters

(MeO) 3 P=O

(MeO) 2 PO 2 H

H 218 O

LiBr

MeOH PhS

O

O

O

BP

P

MeO

O

O

O

BP

O

O

O

O

BP

P

O

O

O

O

BP

O

PhS + PhSMe

+

(MeO) 2 PO 2 Li + MeBr

(MeO) 2 PO 2 + PhSMe

5'

3'

5'

3'

a

b

P = protecting group

Figure 3.46 C O cleavage reactions of triesters with soft nucleophiles. (a) trimethyl phosphate; (b) a phosphate
triester following DNA synthesis