Total synthesis of uridine diphosphate-N-acetylmuramoyl-l-alanine

Article abstract of DOI:10.1016/j.tetasy.2008.09.029

The first total synthesis of uridine diphosphate N-acetylmuramoyl-l-alanine in 13% overall yield is presented. The 11-step synthetic route is based on the synthetic strategy used for the synthesis of uridine diphosphate N-acetylmuramic acid, the MurC ligase substrate. However, an unexpected amide bond cleavage under basic conditions demanded crucial modifications of the final synthetic steps. The total chemical synthesis of MurD ligase substrate provides an excellent alternative to chemoenzymatic synthesis.

Full text of DOI:10.1016/j.tetasy.2008.09.029

Tetrahedron: Asymmetry 19 (2008) 2265–2271
Contents lists available at ScienceDirect
Tetrahedron: Asymmetry
journal homepage: www.elsevier.com/locate/tetasy
Total synthesis of uridine diphosphate-N-acetylmuramoyl-L-alanine
a,b
ˇ a,
ˇ
*
Andrej Babic , Slavko Pecar
a
ˇ
Faculty of Pharmacy, University of Ljubljana, Aškerceva 7, 1000 Ljubljana, Slovenia
b
ˇ
Jozef Stefan Institute, Jamova 39, 1000 Ljubljana, Slovenia
a r t i c l e i n f o
a b s t r a c t
Article history:
The first total synthesis of uridine diphosphate N-acetylmuramoyl-L-alanine in 13% overall yield is pre-
Received 14 August 2008
Accepted 30 September 2008
Available online 22 October 2008
sented. The 11-step synthetic route is based on the synthetic strategy used for the synthesis of uridine
diphosphate N-acetylmuramic acid, the MurC ligase substrate. However, an unexpected amide bond
cleavage under basic conditions demanded crucial modifications of the final synthetic steps. The total
chemical synthesis of MurD ligase substrate provides an excellent alternative to chemoenzymatic
synthesis.
Ó 2008 Elsevier Ltd. All rights reserved.
1. Introduction
successful synthesis of MurD ligase substrate in 11 synthetic
steps.
Antibiotic resistance represents an increasing threat to public
health.1 Multi-drug resistant strains of bacteria are stimulating re-
search into antimicrobial compounds that inhibit the growth of
bacteria by new mechanisms of action.2 The inhibition of essential
targets within the bacteria is a good strategy and the enzymes in-
volved in the biosynthesis of the bacterial peptidoglycan constitute
such a group of targets.3–5 Biological evaluation of inhibitors with
enzymatic assays requires large quantities of commercially
unavailable substrates. These substrates have been obtained from
bacterial extracts,6 by enzymatic,7–9 chemo-enzymatic10,11 or, very
rarely, by total chemical synthesis.12–14 MurD enzyme15,16 is the
second consecutive Mur ligase which almost universally attaches
2. Results and discussion
The first three reaction steps (Scheme 1) were part of the estab-
lished synthesis of UDP-N-acetylmuramic acid. The synthesis
started from N-acetylglucosamine 1, which was protected by benz-
ylation with benzyl alcohol at the anomeric position. Even though
this protecting group was removed later in the synthesis and, the-
oretically, the synthesis could proceed with the
benzyl anomer 2, it was decided that a permanent
figuration of the N-acetylglucosamine system would simplify the
a
/b mixture of
a-anomeric con-
isolation and purification of the correct diastereoisomers after
the stereoselective reactions still to come. Pure benzyl
D
-glutamic acid to the growing peptide chain of the peptidoglycan
a-anomer
precursor and uses UDP-N-acetylmuramoyl-L-alanine 15 as the
nucleotide substrate.
2 was obtained with thermodynamic control of the reaction by
high reaction temperatures in refluxing toluene. The introduction
of a benzylidene protection group to the 40-and 60-hydroxy posi-
tions was carried out according to Gross and Rimpler19 to afford
We have previously reported an improved total synthesis of
UDP-N-acetylmuramic acid,17 the MurC ligase18 nucleotide sub-
strate. Our strategy was designed in such a way that, if needed, it
would allow the total chemical synthesis of other Mur ligase sub-
strates. Both the UDP-N-acetylmuramic acid and UDP-N-acetyl-
protected
a
-N-acetylglucosamine derivative 3 with a free 30-hy-
droxy group. By treating 3 with racemic 2-chloropropionic acid,
the Williamson stereoselective ether synthesis proceeded in good
yield, to afford sodium salt 4 as a mixture of two diastereoisomers.
1H NMR was used to determine the stereoselectivity of the reac-
tion. Integration of the benzylidene protons at 5.66 and 5.55 ppm
for the muramic and isomuramic acid derivatives, respectively,
showed a diastereoisomeric ratio of 74:26 in favour of the mura-
mic acid derivative 4 with the (R)-configuration of the lactoyl
side-chain. This marks the dividing point of the synthetic pathways
to substrates of different Mur ligases.
muramoyl-L-alanine 15 are highly functionalized structurally
complex natural molecules with several stereogenic centers origi-
nating from enantiopure natural carbohydrates. However, over the
course of the total synthesis several key stereogenic centers have
to be introduced by stereoselective steps and any reactions which
could lead to racemization must be avoided at all cost.
Herein, we report the synthetic route to UDP-N-acetylmura-
moyl-L-alanine 15 with important modifications to our previous
strategy towards Mur ligases’ substrates which allowed the
When the sodium salt 4 was reacted with methyliodide in DMF,
this gave methyl ester 5 in quantitative yield and led towards UDP-
N-acetylmuramic acid as previously reported.17 On the other hand,
* Corresponding author. Tel.: +386 1 476 9500; fax: +386 1 425 8031.
when the methyl ester of L-alanine was coupled with 4, this gave
ˇ
E-mail address: andrej.babic@ffa.uni-lj.si (A. Babic).
0957-4166/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetasy.2008.09.029

ˇ
ˇ
2266
A. Babic, S. Pecar / Tetrahedron: Asymmetry 19 (2008) 2265–2271
OH
OH
Ph
O
O
O
O
b
a
O
HO
HO
HO
HO
HO
Ac NH
Ac NH
AcNH
OH
O
Ph
O
Ph
2
3
1
Ph
O
O
O
O
c
Ac
NH
O
Ph
O-
Na+
O
4
d
e
Ph
O
O
O
O
Ph
O
MurD ligase
substrate
MurC ligase
substrate
O
O
O
Ac
O
NH
Ac
NH
O
Ph
O
Ph
NH
O
O
OMe
OMe
5
6
Scheme 1. Reagents and conditions: (a) BnOH, PTSA, toluene, reflux, 3 h, 80%; (b) (CH3CH2O)3CH, PhCHO, dioxane, DMF, rt, 24 h, 76%; (c) 2-chloro-propionic acid, NaH,
dioxane, 45 °C, 16 h; (d) (i) MeI, DMF, rt, 4 h; (ii) diastereoisomer separation, 75% (two steps); (e) (i) L-Ala-OMe, EDC, Hobt, TEA, CH2Cl2, rt, 12 h; (ii) diastereoisomer
separation, 65% (two steps).
amide 6, an intermediate towards the MurD substrate (Scheme 1).
Careful temperature control and the appropriate addition of
reagents resulted in no racemization being observed at this step,
while the yield remained good at 65% for the two steps.20 The
isomuramic diastereoisomers of 5 and 6 with an (S)-configuration
in the lactoyl group were removed by chromatography in both
cases. Next, the benzylidene group was removed by acidolysis in
aqueous 60% acetic acid and was replaced by a base-labile and
hydrogenolysis resistant acetate ester protecting group, to afford
the diacetyl intermediate 7 (Scheme 2). As expected, the overall
yield for both reaction steps was high at 89%. This was followed
by removal of the anomeric benzyl protecting group by hydrogen-
olysis in 98% yield.
duction to the anomer position with subsequent oxidation,23–25
this offered a significant advantage in simplicity, yield and stereo-
selectivity of the reaction.
Therefore, hydrogenation in THF, with PtO2 used as a catalyst,
allowed a straightforward conversion to unprotected phosphate
10, which was quenched immediately by 1 M LiOH to stabilize
the free
ter protecting groups, according to the existing synthetic strategy
(Scheme 2). Surprisingly, the reaction mostly afforded -1-phos-
pho-N-acetylmuramic acid 11. Obviously, the amide bond between
a-phosphate group and remove the acetate and methyl es-
a
the muramic acid and
conditions.
L-alanine moieties was cleaved under these
We reasoned that steric and electronic effects brought about the
otherwise unexpected cleavage of the amide bond since the amide
bond of the N-acetylamide moiety remained intact under the same
reaction conditions. The use of more diluted lithium hydroxide and
THF mixtures did not improve the outcome of this reaction. There-
fore, we assumed that the THF/water solvent system played an
important role in this unwanted cleavage. However THF, present
from the previous reaction step, could not be removed because of
the instability of the free phosphate 10, so changes in the deprotec-
tion and the final diphosphate coupling steps had to be introduced.
For this reason, diphenylphosphate 9 was not globally deprotec-
ted but stabilized by the direct addition of triethylamine to the
hydrogenation reaction mixture. After purification on a short LH-
20 Sephadex column, with methanol as eluent, the triethylamine
salt of 12 was obtained as a crystalline solid. Phosphate 12 was
then subjected to a variety of deprotection conditions, with the
intention of finding conditions that would remove all the protect-
ing groups in quantitative yield but not cause cleavage of the labile
amide bond. When the reaction was carried out in 50 mM aqueous
sodium hydroxide and the pH of the reaction mixture was allowed
It is noteworthy that this seemingly facile reaction proceeds
very slowly, and with low yield, if the wrong reaction solvent is
used.21 In our hands the conversion of 7 to hemiacetal 8 was fast,
reproducible and proceeded with quantitative yields only when
appropriate Pd/C and THF were used as solvents (Scheme 2). The
introduction of an
a-phosphate was achieved according to the
modified procedure of Sabesan and Neira.22 The reaction was
kinetically controlled and carried out at ꢀ25 °C in dry dichloro-
methane with the addition of 4-pyrrolidinopyridine as the catalyst,
followed by slow addition of diphenylchlorophosphate. This affor-
ded just the
a-anomer of 9 in 72% yield, as evidenced from
300 MHz 1H NMR spectrum which showed a dd at 6.75 with the
coupling constants of 5.4 and 3.1 Hz characteristic of the anomeric
proton of
a-phosphates. Diphenylphosphate 9 had to be purified
and handled with care because of its inherent instability. The
diphenyl phosphate group was utilized because it could be intro-
duced with a single reaction step and allowed convenient and high
yielding deprotection under hydrogenation conditions. Compared
to procedures which involve the one-pot dibenzyl phosphite intro-

ˇ
ˇ
A. Babic, S. Pecar / Tetrahedron: Asymmetry 19 (2008) 2265–2271
2267
AcO
AcO
AcO
AcO
Ph
O
O
O
O
O
O
O
b
O
a
AcNH
AcNH
AcNH
O
Ph
OH
O
Ph
NH
NH
O
O
NH
O
O
O
O
OMe
OMe
OMe
8
7
6
c
AcO
AcO
AcO
AcO
O
O
HO
HO
O
e
O
d
O
O
P
O
Ph
O
O
Ac NH
Ac NH
P
O
AcNH
O
O
OH
P
Li+
NH
O
O
O
O
NH
HO
O
O
Li+
Li+
O
O
O
Ph
O
OMe
OMe
11
9
10
f
AcO
AcO
HO
O
O
HO
g
O
O
O
O
P
Ac
NH
OH
P
Ac NH O
O
O
Na+
O
NH
O
NH
O
O
O
Na+
O
+
NH
OMe
12
Na+
13
O
Scheme 2. Reagents and conditions: (a) (i) 60% AcOH, 90 °C, 1 h; (ii) Ac2O, Py, rt, 12 h, 89%; (b) H2, Pd/C (10%), THF, rt, 24 h, 98%; (c) ClPO(OPh)2 4-pyrrolidinopyridine, CH2Cl2,
ꢀ25 °C, 24 h, 72%; (d) H2, PtO2, THF, rt, 24 h; (e) 1 M LiOH, THF/H2O, rt, 24 h; (f) TEA (3 equiv), 92%; (g) 50 mM aqueous NaOH, rt, 4 h, quant.
to drop as the reaction progressed, the target compound
a
-1-phos-
graphy did not allow for the exchange of the cations, so the
protected diphosphate 14 was obtained in the form of the 4-mor-
pholino-N,N0-dicyclohexylcarboxamidine salt. This presented no
major problem since the cations could be conveniently exchanged
after the final reaction step. As already described for the mono-
phosphate 12, 50 mM sodium hydroxide was used for deprotection
of the diphosphate 14 in the final step. Since the sodium hydroxide
was diluted enough to allow the pH to drop as the reaction pro-
ceeded, no amide bond hydrolysis occurred. After the straightfor-
ward desalting Sephadex G-10 column, which removed the
unwanted 4-morpholino-N,N0-dicyclohexylcarboxamidine cations
pho-N-acetylmuramoyl- -alanine 13 could be obtained without
L
any side products and was stable, even if the reaction time was
prolonged (Scheme 2).
In spite of this encouraging result, it was decided that the final
deprotection of the acetate and methyl ester groups would be
accomplished at the very end of the total synthesis for two reasons.
It has been reported that the coupling reaction with uridine-50-
monophospho morpholidate proceeds much more efficiently if
the coupled sugar-phosphate is still protected26 and, furthermore,
if the obtained protected diphosphate could be purified by flash
chromatography, then the demanding final purification by gel fil-
the trisodium salt of UDP-N-acetylmuramoyl-L-alanine 15 was ob-
tration could be omitted. In view of this, the protected
pho-N-acetylmuramoyl- -alanine was successfully coupled with
uridine-50-monophosphomorpholidate to afford protected UDP-
N-acetylmuramoyl- -alanine 14 without the addition of tetrazole
as catalyst27 (Scheme 3). Even though no catalyst was used, the
a
-1-phos-
tained in very high yield.
L
3. Conclusion
L
In conclusion, we have presented the first total chemical syn-
thesis of UDP-N-acetylmuramoyl- -alanine. The synthetic strategy
protected
a-phosphate 12 was reacted with equimolar amounts
of uridine-50-monophospho morpholidate, 61% yield was achieved
for this demanding coupling reaction.
L
for UDP-N-acetylmuramic acid was employed until an unexpected
hydrolysis of the amide bond occurred at the global deprotection
step. This problem was successfully solved by reversing the order
of diphosphate coupling and the final deprotection. The latter
was carried out as the last step of the synthesis under controlled
and increasingly milder basic conditions. The synthetic path dis-
As anticipated, the use of protected
a-1-phosphate 12 also
made a big difference to the purification procedure. The crude
diphosphate 14 could be effectively purified by flash chromatogra-
phy in a fashion similar to that described by Dinev et al. for uridine
diphospho(13C6)glucose.28 However, in our hands, flash chromato-

ˇ ˇ
A. Babic, S. Pecar / Tetrahedron: Asymmetry 19 (2008) 2265–2271
2268
O
AcO
AcO
AcO
AcO
O
N
O
O
NH
O
O
O
P
O
a
O
P
AcNH
OH
P
N
O
Ac
O
O
NH
O
O
O
O
+
NH2
NH
O
O
OH
NH
O
O
N
O
+
NH
OH OH
OMe
12
OMe
14
b
O
N
HO
HO
O
NH
O
O
O
P
O
Ac
O
NH
P
O
O
O
O
NH
O
O
O
Na+
Na+
OH OH
Na+
O
15
Scheme 3. Reagents and conditions: (a) uridine-5-phosphomorpholidate, 4 Å molecular sieves, DMF, 60 °C, 20 h, 61%; (b) (i) 50 mM aqueous NaOH, rt, 4 h; (ii) Sephadex G-10
desalting column, 95%.
covered, with an overall yield of 13% provides a good alternative to
the chemoenzymatic synthesis, especially if scale-up is required.
This is currently in progress in our laboratory.
colourless crystals (28.1 g, 90.4 mmol, 80% yield). Mp 180–
182 °C. IR (KBr, cmꢀ1): 3285, 2931, 1630, 1552, 1376, 1027, 734,
695. ½a 2D0
ꢂ
¼ þ265:6 (c 0.08, DMF). 1H NMR (DMSO-d6, 300 MHz):
d (ppm) = 7.79 (d, 1H, J = 8.1 Hz, NH), 7.39–7.26 (m, 5H, Ph–H),
4.99 (d, 1H, J = 5.6 Hz, OH-4), 4.72–4.70 (m, 2H, H-1, OH-3), 4.67
(d, 1H, Jgem = 12.6 Hz, CH2a–Ph), 4.52 (t, 1H, J = 5.8 Hz, OH-6),
4.42 (d, 1H, Jgem = 12.6 Hz, CH2b–Ph), 3.72–3.62 (m, 2H, H-2,
H-6), 3.57–3.43 (m, 3H, H-3, H-60, H-5), 3.16 (ddd, 1H, J = 9.1,
9.1, 5.6 Hz, H-4), 1.84 (s, 3H, COCH3). 13C NMR (DMSO-d6,
75 MHz): d (ppm) 169.4, 137.9, 128.1, 127.5, 127.4, 95.9, 73.1,
70.9, 70.6, 67.7, 60.8, 53.7, 22.5. LRMS (FAB), m/z = 312 (M+H)+,
(ESI) m/z = 334.1 (M+Na)+. HRMS (ESI), m/z calcd for C15H21NO6Na
334.1267 (M+Na)+, found 334.1270.
4. Experimental
All reactions were carried out under an argon atmosphere using
anhydrous solvents, unless indicated otherwise. Chemicals from
Sigma–Aldrich, Fluka and Merck (10% Pd/C) were used without fur-
ther purification. Analytical TLC was performed on Merck Silica Gel
(60F 254) plates (0.25 mm) and components visualized with ultra-
violet light and stained with 20% sulfuric acid in ethanol, rhoda-
mine G6, 2,4-dinitrophenylhydrazine, bromocresol green and
ninhydrin. Flash chromatography was performed using Merck Sil-
ica Gel (0.040–0.063 mm). Gel filtration was performed using
Sephadex G-10 and LH-20 stationary phases and methanol or
bidistilled water as eluents. 1H, 13C, 31P, DEPT-135, gradient COSY
and gradient HSQC NMR spectra were recorded on a Bruker
AVANCE DPX300 spectrometer at 302 K in CDCl3, DMSO-d6, D2O
and MeOH–d4 solutions using TMS or residual undeuterated sol-
vent as the internal standard. Microanalyses were performed on
a Perkin–Elmer C, H, N analyzer 240 C. Optical rotation was mea-
sured using Perkin–Elmer 241 MC polarimeter at 589 nm.
Reverse-phase HPLC analysis was performed using Agilent 1100
Series system and Phenomenex Luna C18 5u (250 ꢁ 4.60 mm) col-
umn. Mass spectra were obtained using a VG-Analytical Autospec
Q and Q-TOF Premier mass spectrometers.
4.2. Benzyl 4,6-O-benzylidene-2-acetylamido-2-deoxy-a-D-
glucopyranoside 3
Compound 3 was synthesized according to the literature proce-
dure.11 Compound 2 (15 g, 48.2 mmol) was dried using absolute
ethanol and toluene co-evaporation under reduced pressure. The
dry solid was suspended in 50 mL of dry DMF, 50 mL anhydrous
dioxane, 25 mL triethylorthoformate and 20 mL of freshly distilled
benzaldehyde. p-Toluenesulfonic acid monohydrate (1.2 g,
6.31 mmol) was added and the reaction mixture stirred at ambient
temperature. After 24 h, the reaction mixture was poured into an
Erlenmeyer flask containing 300 mL of ether. The suspension was
stirred at 0 °C and after 1 h the colourless precipitate was filtered
off, washed with an additional 100 mL of ether and dried in vacuo.
Crystallization from absolute ethanol afforded a colourless solid
(14.6 g, 36.6 mmol, 76%). Mp >240 °C. IR (KBr, cmꢀ1): 3300, 3066,
4.1. Benzyl-2-deoxy-2-acetylamido-a-D-glucopyranoside 2
N-Acetyl-glucosamine 1 (25 g, 113 mmol) and p-toluenesul-
fonic acid monohydrate (1.9 g, 10 mmol) were suspended in
300 mL of toluene and 180 mL of benzyl alcohol. The reaction
mixture was refluxed in a Dean–Stark apparatus with water
removal by azeotrope mixture. After 3 h, the reaction mixture
was cooled to ambient temperature and sodium bicarbonate
(1.26 g, 15 mmol) dissolved in water was added. Toluene was
removed under reduced pressure and ether–hexane (2:1) mixture
(900 mL) was added to the oily residue and stirred vigorously for
3 h. The amorphous precipitate was filtered off, washed with
ether and the crude product recrystallized from ethanol to yield
2966, 2868, 1650, 1552, 1372, 1013, 928, 727, 592. ½a D20
¼ þ49:3
ꢂ
(c 0.20, DMF). 1H NMR (CDCl3, 300 MHz): d (ppm) = 7.52–7.26
(m, 10H, Ph–H), 5.79 (d, 1H, J = 8.7 Hz, NH), 5.57 (s, 1H, CH–Ph),
4.94 (d, 1H, J = 3.8 Hz, H-1), 4.78 (d, 1H, Jgem = 11.8 Hz, CH2b–Ph),
4.50 (d, 1H, Jgem = 11.8 Hz, CH2a–Ph), 4.28–4.20 (m, 2H, H-2, H-6),
3.96 (ddd, 1H, J = 9.2, 9.2, 3.0 Hz, H-5), 3.86 (dd, 1H, J = 9.5 Hz,
4.6 Hz, H-3), 3.79–3.70 (m, 1H, H-60), 3.60 (app t, 1H, J = 9.2 Hz,
H-4), 2.96 (d, 1H, J = 3.3 Hz, OH-3), 1.99 (s, 3H, CH3CO). 13C NMR
(CDCl3, 75 MHz): d (ppm) 171.3, 137.0, 136.8, 129.2, 128.7, 128.3,
128.3, 128.1, 126.3, 101.9, 97.2, 82.1, 70.6, 70.1, 68.8, 62.8, 54.0,
23.2. LRMS (FAB), m/z = 400 (M+H)+, (ESI) m/z = 422.2 (M+Na)+.

ˇ
ˇ
A. Babic, S. Pecar / Tetrahedron: Asymmetry 19 (2008) 2265–2271
2269
HRMS (ESI), m/z calcd for C22H25NO6Na 422.1580 (M+Na)+, found
422.1578.
The colourless foam was dried for several hours in vacuo and used
in the next reaction step. The crude product was dissolved in pyr-
idine (40 mL), flushed with argon and cooled to 0 °C. While being
stirred, acetic acid anhydride (5.0 mL, 45.4 mmol) was added drop-
wise. The reaction mixture was allowed to warm to ambient tem-
perature and stirred overnight. Methanol (10 mL) was added
slowly to quench the excess acetic acid anhydride and the solvents
were evaporated under reduced pressure. Toluene co-evaporation
was used to remove traces of pyridine. The crude product was puri-
fied using flash chromatography (dichloromethane–metha-
nol = 15:1). A colourless solid (1.78 g, 3.22 mmol, 89% yield) was
obtained. Mp 148–149 °C. IR (KBr, cmꢀ1): 2929, 1745, 1658,
1563, 1548, 1535, 1451, 1376, 1240, 1123, 1044, 668.
4.3. (S)-Methyl 2-(2-(R)-(2-acetamido-1-O-benzyl-4,6-O-
benzylidene-2-deoxy-3-a-D-glucopyranosyloxy)
propanamido)propanoate 6
Compound 3 (10.0 g, 25.0 mmol) was suspended in 100 mL of
benzene and the solvent was evaporated under reduced pressure
to remove traces of water. The colourless solid was dissolved in
100 mL of dry dioxane and flushed with argon. Sodium hydride
(4.20 g, 175 mmol) was slowly added at room temperature. The
reaction mixture thickened significantly on heating at 50 °C for
1 h under argon atmosphere. The temperature was then lowered
to 45 °C and racemic 2-chloro-propionic acid (8.0 g, 74 mmol) dis-
solved in dry dioxane (20 mL) was slowly added dropwise. After
the addition, the mixture was heated at 45 °C for 16 h, when the
TLC analysis showed total consumption of the starting material.
The reaction mixture was cooled to ambient temperature and
quenched carefully with water. The solvents were removed under
reduced pressure and, after the addition of 100 mL of brine, the so-
dium salt was left overnight to precipitate at 0 °C. The amorphous
precipitate was suction filtered and dried at 50 °C. A slightly
brownish solid (15.0 g, 30.4 mmol, 125%) was obtained and used
in the next reaction step without purification. The crude sodium
salt was suspended in dry dichloromethane (500 mL) and cooled
½
a 2D0
ꢂ
¼ þ77:2 (c 0.18, DMF). 1H NMR (CDCl3, 300 MHz): d (ppm)
7.24–7.27 (m, 5H, Ph–H), 6.87 (d, 1H, J = 7.2 Hz, NH), 6.95 (d, 1H,
J = 9.5 Hz, NH), 5.08 (app t, 1H, J = 9.7 Hz, H-4), 4.97 (d, 1H,
J = 3.6 Hz, H-1), 4.70 (d, 1H, Jgem = 11.7 Hz, CH2a–Ph), 4.50 (d, 1H,
Jgem = 11.7 Hz, CH2b–Ph), 4.43–4.33 (m, 2H, NHCHCH3, H-2), 4.20
(dd, 1H, J = 12.4, 4.6 Hz, H-6), 4.07 (dd, 1H, J = 12.4, 2.4 Hz, H-60),
3.98 (q, 1H, J = 6.7 Hz, OCHCH3), 3.91 (ddd, 1H, J = 10.2, 4.4,
2.4 Hz, H-5), 3.71 (s, 3H, COOCH3), 3.66 (app t, 1H, J = 9.7 Hz,
H-3), 2.11 (s, 3H, COCH3), 2.08 (s, 3H, COCH3), 1.89 (s, 3H,
NHCOCH3), 1.44 (d, 3H, J = 7.2 Hz, NHCHCH3), 1.32 (d, 3H,
J = 6.7 Hz, OCHCH3). 13C NMR (CDCl3, 75 MHz): d (ppm) 172.8,
172.1, 170.7, 170.1, 169.3, 136.6, 128.7, 128.4, 128.3, 97.0, 78.7,
78.4, 70.2, 69.5, 68.5, 62.1, 53.0, 52.3, 48.0, 23.3, 20.8, 20.8, 18.7,
17.4. LRMS (ESI), m/z = 553.2 (M+H)+, 575.2 (M+Na)+, HRMS (ESI),
m/z calcd for C26H37N2O11 553.2397 (M+H)+, found 553.2400.
Microanalysis calcd for C26H36N2O11 (%): C, 56.51; H, 6.57; N,
5.07; found: C, 56.15; H, 6.72; N, 4.94.
to 0 °C in an ice-bath under argon atmosphere. L-Alanine methyl
ester hydrochloride (6.36 g, 45.6 mmol), 1-hydroxybenzotriazole
(4.53 mg, 33.5 mmol) and finally 1-ethyl-3-(3-dimethylaminopro-
pyl)-carbodiimide (6.42 g, 33.5 mmol) were added to the stirred
suspension. Triethylamine was used to adjust the pH value to 8
and the mixture was allowed to warm slowly to ambient temper-
ature. After stirring overnight the suspension was diluted with
dichloromethane (200 mL) and washed with sodium bicarbonate,
dilute citric acid and finally brine. The organic phase was dried
over sodium sulfate and concentrated to dryness under reduced
pressure. The crude product was purified using flash chromatogra-
phy (dichloromethane–ethyl acetate–hexane = 3:3:1). A colourless
solid (9.1 g, 16.4 mmol, 65% yield) was obtained. Mp 220–222 °C.
IR (KBr, cmꢀ1): 3090, 2931, 2870, 1742, 1657, 1554, 1499, 1451,
1372, 1321, 1214, 1156, 1124, 1090, 1055, 1028, 1006, 973, 733,
4.5. (S)-Methyl 2-(2-(R)-(2-acetamido-4,6-di-O-acetyl-2-deoxy-
3-D
-glucopyranosyloxy)propanamido) propanoate 8
Compound 7 (1.60 g, 2.89 mmol) was dissolved in freshly dis-
tilled dry THF (200 mL). Palladium on charcoal (700 mg, 10%)
was added after the solution was flushed with argon for 10 min.
The reaction mixture was first flushed with hydrogen for 30 min
and then stirred under positive hydrogen pressure at ambient tem-
perature. After 24 h, the catalyst was filtered off and washed with a
small amount of THF. The solvent was evaporated under reduced
pressure giving a colourless foam (1.32 g, 2.86 mmol, 98% yield).
The product was used without purification in the next reaction
step. Mp 84–87 °C. IR (KBr, cmꢀ1): 2957, 1746, 1659, 1547, 1452,
695. ½a 2D0
ꢂ
¼ þ44:6 (c 0.21, DMF). 1H NMR (CDCl3, 300 MHz): d
(ppm) 7.90–7.26 (m, 10 H, Ph–H), 6.90 (d, 1H, J = 7.2 Hz, NH),
6.13 (d, 1H, J = 7.2 Hz, NH), 5.57 (s, 1H, CH–Ph), 4.97 (d, 1H,
J = 3.7 Hz, H-1), 4.72 (d, 1H, Jgem = 11.8 Hz, CH2a–Ph), 4.51–4.43
(m, 2H, CH2a–Ph, NHCHCH3), 4.33–4.29 (m, 1H, H-2), 4.24 (dd,
1H, J = 9.9, 4.5 Hz, H-6), 4.11 (q, 1H, J = 6.7 Hz, OCHCH3), 3.87
(ddd, 1H, J = 9.3, 9.0, 4.5 Hz, H-5), 3.80–3.67 (m, 6H, H-60, H-3,
COOCH3, H-4), 1.93 (s, 3H, COCH3), 1.42 (d, 3H, J = 7.6 Hz,
NHCHCH3), 1.39 (d, 3H, J = 6.7 Hz, OCHCH3). 13C NMR (CDCl3,
75 MHz): d (ppm) 173.0, 172.9, 170.3, 137.1, 136.8, 129.0, 128.7,
128.3, 128.2, 125.9, 101.5, 97.6, 81.7, 78.4, 77.7, 77.2, 70.2, 68.8,
63.2, 53.1, 52.4, 48.0, 23.4, 19.4, 17.9. LRMS (FAB), m/z = 557
(M+H)+, (ESI) m/z = 557.3 (M+H)+, 579.2 (M+Na)+. HRMS (ESI), m/
z calcd for C29H36N2O9Na 579.2319 (M+H)+, found 579.2315.
Microanalysis calcd for C29H36N2O9 (%): C, 62.58; H, 6.52; N,
5.03; found: C, 62.85; H, 6.73; N, 5.03.
1376, 1242, 1121, 1043, 601. ½a D20
ꢂ
¼ þ40:3 (c 0.12, MeOH). 1H
NMR (CDCl3, 300 MHz): d (ppm) 6.97 (d, 1H, J = 7.2 Hz, NH), 6.90
(d, 1H, J = 8.9 Hz, NH), 5.26 (s, 1H, OH), 5.14 (d, 1H, J = 3.2 Hz, H-
1), 4.99 (app t, 1H, J = 9.7 Hz, H-4), 4.44–4.30 (m, 1H, NHCHCH3),
4.19–4.00 (m, 5H, H-2, H-6, H-5, OCHCH3, H-60), 3.72 (app t, 1H,
J = 9.6 Hz, H-3), 3.68 (s, 3H, COOCH3), 2.05 (s, 3H, COCH3), 2.04 (s,
3H, COCH3), 1.92 (s, 3H, NHCOCH3), 1.40 (d, 3H, J = 7.2 Hz,
NHCHCH3), 1.27 (d, 3H, J = 6.6 Hz, OCHCH3). 13C NMR (CDCl3,
75 MHz): d (ppm) 172.8, 172.6, 171.0, 170.9, 169.5, 91.7, 78.0,
77.3, 69.9, 67.8, 62.4, 53.7, 52.4, 48.0, 23.2, 20.8, 20.7, 18.6, 17.4.
LRMS (ESI), m/z = 463.2 (M+H)+, 445.2 (MꢀH2O)+, 485.2 (M+Na)+,
HRMS (ESI), m/z calcd for C19H31N2O11 463.1928 (M+H)+, found
463.1949. Microanalysis calcd for C19H30N2O11(%): C, 49.35; H,
6.54; N, 6.06; found: C, 48.99; H, 6.69; N, 5.80.
4.4. (S)-Methyl 2-(2-(R)-(2-acetamido-4,6-di-O-acetyl-1-O-
benzyl-2-deoxy-3-a-D-glucopyranosyloxy)
propanamido)propanoate 7
4.6. (S)-Methyl 2-(2-(R)-(2-acetamido-2-deoxy-4,6-di-O-acetyl-
1-O-diphenoxyphosphoryl-3-a-D-glucopyranosyloxy)-
propanamido)propanoate 9
Compound 6 (2.00 g, 3.60 mmol) was suspended in aqueous
acetic acid (50 mL, 60%) and heated at 90 °C. After 1 h, the solvents
were evaporated under reduced pressure. Repetitive toluene co-
evaporation was used to remove traces of water and acetic acid.
Compound 8 (335 mg, 0.73 mmol) and 4-pyrrolidinopyridine
(536 mg, 3.62 mmol) were dissolved in dry dichloromethane

ˇ ˇ
A. Babic, S. Pecar / Tetrahedron: Asymmetry 19 (2008) 2265–2271
2270
(30 mL). The reaction flask was flushed with argon and cooled to
4.8. 4-Morpholino-N,N0-dicyclohexylcarboxamidine salt of (S)-
methyl 2-(2-(R)-(2-acetamido-2-deoxy-4,6-di-O-acetyl-1-O-
ꢀ25 °C. After being stirred for 20 min, diphenyl chlorophosphate
(450
lL, 2.16 mmol) was added dropwise using a syringe. After
(uridine-50-diphosphoryl)-3-
propanamido)propanoate 14
a-D-glucopyranosyloxy)-
24 h, at ꢀ25 °C under an argon atmosphere the reaction mixture
was diluted with dichloromethane (30 mL) and washed consecu-
tively with cold citric acid (5%), saturated sodium bicarbonate solu-
tion and brine. The organic phase was dried over sodium sulfate
and the solvent evaporated under reduced pressure at a tempera-
ture not exceeding 25 °C. The crude product was purified using
gradient flash chromatography (ethyl acetate–hexane = 50:50 to
100:0) yielding a transparent oil which turned to a colourless foam
(362 mg, 0.52 mmol, 72% yield) after treatment with ether and dry-
ing in vacuo. IR (KBr, cmꢀ1): 3072, 2360, 2340, 1748, 1662, 1592,
1498, 1455, 1373, 1212, 1164, 1111, 1045, 964, 922, 778, 691,
Compound 12 (140 mg, 0.22 mmol) was suspended in dry benz-
ene (10 mL) and the solvent was evaporated. After the starting
material was dried in vacuo for 1 h, dry DMF stored over molecular
sieves, the 4-morpholine-N,N0-dicyclohexylcarboxamidine salt of
uridine monophosphate morpholidate (160 mg, 0.23 mmol) and
4 Å molecular sieves were added. The reaction mixture was flushed
with argon and first stirred at room temperature for 30 min and
then heated at 60 °C. After 24 h, water (1 mL) was added and the
reaction was stirred for an additional 24 h. The solvents were evap-
orated under reduced pressure at a temperature not exceeding
30 °C. The crude product was purified by gradient flash chromato-
graphy (5–20% of water in acetonitrile). A colourless solid was
obtained (175 mg, 0.13 mmol, 61% yield). Mp 164–165 °C. IR
(KBr, cmꢀ1): 2936, 2858, 1745, 1680, 1620, 1564, 1452, 1377,
617, 539. ½a 2D0
ꢂ
¼ þ61:4 (c 0.22, DMF). 1H NMR (CDCl3, 300 MHz):
d (ppm) 7.39–7.20 (m, 10H, Ph–H), 6.79 (d, 1H, J = 8.1 Hz, NH),
6.72 (d, 1H, J = 7.2 Hz, NH), 6.75 (dd, 1H, J = 5.4, 3.1 Hz, H-1), 5.14
(app t, 1H, J = 9.6 Hz, H-4), 4.51–4.41 (m, 1H, NHCHCH3), 4.41–
4.34 (m, 1H, H-2), 4.13 (dd, 1H, J = 12.4, 3.9 Hz, H-6), 4.05 (q, 1H,
J = 6.7 Hz, OCHCH3), 4.02–3.97 (m, 1H, H-5), 3.91 (dd, 1H, J = 12.4,
2.9 Hz, H-60), 3.74 –3.69 (m, 4H, COOCH3, H-3), 2.09 (s, 3H, COCH3),
2.00 (s, 3H, COCH3), 1.78 (s, 3H, NHCOCH3), 1.43 (d, 3H, J = 7.2 Hz,
NHCHCH3), 1.32 (d, 3H, J = 6.9 Hz, OCHCH3). 13C NMR (CDCl3,
75 MHz): d (ppm) 172.7, 172.1, 170.7, 170.6, 169.0, 150.3, 150.2,
123.0, 125.8, 125.7, 120.1, 120.0, 119.9, 119.8, 97.8, 78.1, 76.4,
70.5, 69.0, 61.3, 53.3, 53.2, 52.4, 48.0, 22.9, 20.7, 20.6, 18.9, 17.6.
31P NMR (CDCl3, 121 MHz): d (ppm) ꢀ13.20 (s, 1P). LRMS (ESI),
m/z = 695.3 (M+H)+, 717.2 (M+Na)+, 733.2 (M+K)+, HRMS (ESI), m/
z calcd for C31H39N2O14PNa 717.2037 (M+Na)+, found 717.2045.
Microanalysis calcd for C31H39N2O14P (%): C, 53.60; H, 5.66; N,
4.03; found: C, 53.71; H, 5.80; N, 4.22.
1243, 1116, 1040, 1002, 929, 520. ½a D20
¼ þ105:3 (c 0.08, DMF).
ꢂ
1H NMR (MeOH-d4, 300 MHz): d (ppm) 8.05 (d, 1H, J = 8.2 Hz,
CH), 5.99 (d, 1H, J = 4.9 Hz, H-1r), 5.87 (d, 1H, J = 8.3 Hz, CH), 5.60
(dd, 1H, J = 7.4, 2.9 Hz, H-1g), 5.10 (app t, 1H, J = 9.6 Hz, H-4g),
4.38–4.17 (m, 10H, H-2r, H-3r, H-4r, H-5r, H-50 H-2g, H-5g, H-6g,
r
H-60 , OCHCH3, NHCHCH3), 3.98 (app t, 1H, J = 9.8 Hz, H-3g), 3.77
g
(app t, 4H, J = 4.8 Hz, 2CH2O), 3.73 (s, 3H, COOCH3), 3.44 (app t,
4H, J = 4.8 Hz, 2CH2N), 3.38–3.33 (m, 2H, 2CH), 2.11 (s, 3H, COCH3),
2.07 (s, 3H, COCH3), 2.03 (s, 3H, NHCOCH3), 1.98–1.69 (m, 10H,
5CH2), 1.44 (d, 3H, J = 7.3 Hz, NHCHCH3), 1.32 (d, 3H, J = 6.6 Hz,
OCHCH3), 1.51–1.15 (m, 10H, 5CH2). 13C NMR (MeOH-d4,
75 MHz): d (ppm) 174.7, 174.3, 174.00, 172.5, 171.6, 166.0,
159.3, 152.6, 142.6, 103.3, 96.2, 90.0, 85.1, 79.1, 78.7, 75.6, 71.2,
70.5, 70.3, 67.3, 66.2, 63.2, 56.0, 55.1, 52.8, 49.8, 34.4, 26.3, 26.1,
23.2, 21.1, 20.8, 19.2, 17.4. 31P NMR (MeOH-d4, 121 MHz): d
(ppm) ꢀ9.8 (d, 1P, J = 20.0 Hz), ꢀ12.2 (d, 1P, J = 20.0 Hz). LRMS
(ESIꢀ), m/z = 847.2 (MꢀH)ꢀ; (ESI+), m/z = 294.3 (M+H)+. HRMS
(ESIꢀ), m/z calcd for C28H41N4O22P2 847.1688 (MꢀH)ꢀ, found
4.7. Triethylammonium salt of (S)-methyl 2-(2-(R)-(2-
acetamido-2-deoxy-4,6-di-O-acetyl-1-O-phosphoryl-3-a-D-
glucopyranosyloxy)propanamido)propanoate 12
Compound 9 (300 mg, 0.44 mmol) was dissolved in freshly dis-
tilled dry THF (50 mL). Platinum oxide (100 mg) was added after
the solution was flushed with argon for 20 min. The reaction mix-
ture was stirred vigorously while the hydrogen gas was bubbled
through for 30 min and then left under hydrogen atmosphere at
847.1709. HPLC: Column C18 Phenomenex Luna 5l; mobile phase:
20% acetonitrile, 80% aqueous trifluoroacetic acid (0.01%), flow rate
1.0 mL/min; injection volume: 10
(99.1% @ 254 nm).
lL; retention time: 15.0 min
ambient temperature for 24 h. Triethylamine (250 lL, 1.78 mmol)
4.9. Trisodium (S)-2-(2-(R)-(2-acetamido-2-deoxy-1-O-
(uridine-50-diphosphoryl)-3-
-glucopyranosyloxy)-
propanamido)propanoate 15
was added and the solvent was evaporated under reduced pres-
sure. The crude product was purified on an LH-20 Sephadex col-
umn using methanol as eluent, giving a very hygroscopic solid
(261 mg, 0.41 mmol, 92% yield). IR (KBr, cmꢀ1): 2939, 2677,
2492, 1745, 1662, 1547, 1452, 1377, 1243, 1168, 1114, 1039,
a-D
Compound 14 (73 mg, 0.064 mmol) was dissolved in bi-distilled
water (4.5 mL) and 1 M sodium hydroxide (240 L) was added. The
l
967, 918, 847, 554.
½
a 2D0
ꢂ
¼ þ39:0 (c 0.09, MeOH). 1H NMR
reaction mixture was stirred at ambient temperature and moni-
tored by HPLC. After 4 h, the pH was adjusted to 8 by the addition
of 1 M HCl. After concentration to approx. 1 mL under reduced
pressure at ambient temperature the crude product was purified
by ion-exclusion chromatography on a G-10 Sephadex column
equilibrated with bi-distilled water. The fractions containing the
product were pooled and evaporated under reduced pressure and
dried using toluene-coevaporation yielding a colourless solid
(50 mg, 0.061 mmol, 95% yield). Mp 210–212 °C. IR (KBr, cmꢀ1):
(MeOH–d4, 300 MHz): d (ppm) 5.44 (dd, 1H, J = 6.9, 3.3 Hz, H-1),
5.08 (app t, 1H, J = 9.6 Hz, H-4), 4.35 (q, 1H, J = 9.6 Hz, NHCHCH3),
4.31–4.22 (m, 3H, H-5, H-2, H-6), 4.17 (q, 1H, J = 6.8 Hz, OCHCH3),
4.14–3.91 (dd, 1H, J = 11.9, 2.3 Hz, H-60), 3.89 (app t, 1H, J = 9.8 Hz,
H-3), 3.73 (s, 3H, COOCH3), 3.14 (q, 6H, J = 6.8 Hz, NCH2CH3), 2.12
(s, 3H, COCH3), 2.07 (s, 3H, COCH3), 1.96 (s, 3H, NHCOCH3), 1.44
(d, 3H, J = 7.3 Hz, NHCHCH3), 1.32 (d, 3H, J = 6.8 Hz, OCHCH3),
1.31 (q, 9H, J = 7.3 Hz, NCH2CH3). 13C NMR (MeOH–d4, 75 MHz): d
(ppm) 175.0, 174.3, 173.5, 172.5, 171.6, 95.3, 79.2, 78.8, 70.8,
70.0, 63.2, 55.2, 52.8, 47.7, 23.1, 21.0, 20.8, 19.3, 17.3, 9.2. 31P
NMR (MeOH–d4, 121 MHz): d (ppm) ꢀ0.17 (s, 1P). LRMS (ESIꢀ),
m/z = 541.1 (MꢀH)ꢀ; (ESI+), m/z = 565.1 (M+Na)+, 581.1 (M+K)+.
HRMS (ESI+), m/z calcd for C19H31N2O14PNa 565.1411 (M+Na)+,
3422, 1664, 1406, 1250, 1120, 930. ½a D20
¼ þ40:7 (c 0.04, MeOH).
ꢂ
1H NMR (D2O, 300 MHz, MeOH standard): d (ppm) 8.35 (d, 1H,
J = 9.5 Hz, NH), 7.90 (d, 1H, J = 8.1 Hz, CH), 7.79 (d, 1H, J = 7.1 Hz,
NH), 5.94 (d, 1H, J = 3.6 Hz, H-1r), 5.93 (d, 1H, J = 8.3 Hz, CH),
5.44 (dd, 1H, J = 7.2, 3.2 Hz, H-1g), 4.35–4.30 (m, 2H, H-2r, H-3r),
4.26–4.22 (m, 1H, H-4r), 4.20–4.14 (m, 4H, OCHCH3, H-5r, H-50 ,
found 565.1409. HPLC: Column C18 Phenomenex Luna 5
l; mobile
r
phase: 20% acetonitrile, 80% aqueous trifluoroacetic acid (0.01%),
NHCHCH3), 4.11–4.04 (m, 1H, H-2g), 3.94–3.89 (m, 1H, H-5g),
3.82–3.75 (m, 2H, H-6, H-60) 3.74 (app t, 1H, J = 9.5 Hz H-3g) 3.60
(t, 1H, J = 9.5 Hz, H-4g), 1.97 (s, 3H, NHCOCH3), 1.35 (d, 3H,
flow rate 1.0 mL/min; injection volume: 10
8.2 min (97.6% @ 210 nm).
lL; retention time:

ˇ
ˇ
A. Babic, S. Pecar / Tetrahedron: Asymmetry 19 (2008) 2265–2271
2271
J = 6.6 Hz, OCHCH3), 1.34 (d, 3H, J = 7.3 Hz, NHCHCH3), 13C NMR
(D2O, 75 MHz, MeOH standard): d (ppm) 178.7, 175.2, 174.4,
166.3, 151.9, 141.8, 102.8, 94.7, 88.7, 83.3, 79.7, 78.1, 73.9, 73.1,
69.8, 68.4, 65.2, 60.5, 53.4, 50.0, 22.3, 18.6, 17.2. 31P NMR (D2O,
121 MHz): d (ppm) ꢀ10.1 (d, 1P, J = 20.0 Hz), ꢀ11.9 (d, 1P,
J = 20.0 Hz). LRMS (ESIꢀ), m/z = 749.2 (MꢀH)ꢀ; (ESI+), m/z = 773.1
(M+Na)+, 795.1 (M+2Na)+, 817.1 (M+3Na)+, 839.1 (M+4Na)+. HRMS
(ESIꢀ), m/z calcd for C23H35N4O20P2 749.1320 (MꢀH)ꢀ, found
7. Benson, T. E.; Marquardt, J. L.; Marquardt, A. C.; Etzkorn, F. A.; Walsh, C. T.
Biochemistry 1993, 32, 2024–2030.
8. Reddy, S. G.; Waddell, S. T.; Kuo, D. W.; Wong, K. K.; Pompliano, D. L. J. Am.
Chem. Soc. 1999, 121, 1175–1178.
9. Bouhss, A.; Dementin, S.; van Heijenoort, J.; Parquet, C.; Blanot, D. Methods
Enzymol. 2002, 354, 189–196.
10. Kurosu, M.; Mahapatra, S.; Narayanasamy, P.; Crick, D. C. Tetrahedron Lett.
2007, 48, 799–803.
11. Raymond, J. B.; Price, N. P.; Pavelka, M. S. FEMS Microbiol. Lett. 2003, 229,
83–89.
12. Blanot, D.; Auger, G.; Liger, D.; van Heijenoort, J. Carbohydr. Res. 1994, 252, 107–
115.
749.1335. HPLC: Column C18 Phenomenex Luna 5l; mobile phase:
2% acetonitrile, 98% aqueous trifluoroacetic acid (0.01%), flow rate
13. Dini, C.; Drochon, N.; Ferrari, P.; Aszodi, J. Bioorg. Med. Chem. Lett. 2000, 10,
143–145.
14. Hitchcock, S. A.; Eid, C. N.; Aikins, J. A.; Zia-Ebrahimi, M.; Blaszczak, L. C. J. Am.
Chem. Soc. 1998, 120, 1916–1917.
1.5 mL/min; injection volume: 10
(100% @ 254 nm, 99.2% @ 210 nm).
lL; retention time: 11.8 min
15. Bertrand, J. A.; Auger, G.; Fanchon, E.; Martin, L.; Blanot, D.; van Heijenoort, J.;
Dideberg, O. EMBO J. 1997, 16, 3416–3425.
16. Bertrand, J. A.; Auger, G.; Fanchon, E.; Blanot, D.; Le Beller, D.; van Heijenoort,
J.; Dideberg, O. J. Mol. Biol. 1999, 289, 579–590.
Acknowledgements
This work was supported by the European Union FP6 Integrated
Project EUR-INTAFAR (Project No. LSHM-CT-2004-512138) under
the thematic priority Life Sciences, Genomics and Biotechnology
for Health and the Ministry of Higher Education, Science and Tech-
nology of the Republic of Slovenia. The authors are thankful to Dr.
ˇ
ˇ
17. Babic, A.; Pecar, S. Tetrahedron Lett. 2007, 48, 4403–4405.
18. Mol, C. D.; Brooun, A.; Dougan, D. R.; Hilgers, M. T.; Tari, L. W.; Wijnands, R. A.;
Knuth, M. W.; McRee, D. E.; Swanson, R. V. J. Bacteriol. 2003, 185, 4152–
4162.
19. Gross, P. H.; Rimpler, M. Liebigs Ann. Chem. 1986, 37–45.
20. Pearson, J. A.; Roush, W. R. Handbook of Reagents for Organic Synthesis. In
Activating Agents and Protecting Groups; John Wiley & Sons: New York, 2001, pp
186–188.
21. Osawa, T.; Sinay, P.; Halford, M.; Jeanloz, R. W. Biochemistry 1969, 8, 3369–
3375.
22. Sabesan, S.; Neira, S. Carbohydr. Res. 1992, 223, 169–185.
23. Zamyatina, A.; Gronow, S.; Puchberger, M.; Graziani, A.; Hofinger, A.; Kosma, P.
Carbohydr. Res. 2003, 338, 2571–2589.
ˇ
ˇ
Bogdan Kralj and Dr. Dušan Zigon of Jozef Stefan Institute for the
mass spectra and Dr. Roger H. Pain for critical reading of the
manuscript.
References
24. Graziani, A.; Zamyatina, A.; Kosma, P. Carbohydr. Res. 2004, 339, 147–
151.
25. Narayan, R. S.; VanNieuwenhze, M. S. Eur. J. Org. Chem. 2007, 1399–
1414.
26. Zamyatina, A.; Gronow, S.; Oertelt, C.; Puchberger, M.; Brade, H.; Kosma, P.
Angew. Chem., Int. Ed. 2000, 39, 4150–4153.
27. Wittmann, V.; Wong, C. H. J. Org. Chem. 1997, 62, 2144–2147.
28. Dinev, Z.; Wardak, A. Z.; Brownlee, R. T. C.; Williams, S. Carbohydr. Res. 2006,
341, 1743–1747.
1. Wright, G. D. Nat. Rev. Microbiol. 2007, 5, 175–186.
2. Wright, G. D.; Sutherland, A. D. Trends Mol. Med. 2007, 13, 260–267.
ˇ
3. Barreteau, H.; Kovac, A.; Boniface, A.; Sova, M.; Gobec, S.; Blanot, D. FEMS
Microbiol. Rev. 2008, 32, 168–207.
4. Zoeiby, A.; Sanschagrin, F.; Levesque, R. C. Mol. Microbiol. 2003, 47,
1–12.
5. Katz, A. H.; Caufield, C. E. Curr. Pharm. Des. 2003, 9, 857–866.
6. Michaud, C.; Blanot, D.; Fluoret, B.; van Heijenoort, J. Eur. J. Biochem. 1987, 166,
631–637.