Nitroxyl peptides as catalysts of enantioselective oxidations

Article abstract of DOI:10.1002/1521-3765(20020104)8:1<84::AID-CHEM84>3.0.CO;2-N

The achiral, nitroxyl-containing α-amino acid TOAC (TOAC = 2,2,6,6-tetramethylpiperidine-1-oxyl-4-amino-4-carboxylic acid), in combination with the chiral α-amino acid C^α-methyl valine [(αMe)Val], was used to prepare short peptides (from di- to hexa-) that induced the enantioselective oxidation of racemic 1-phenylethanol to acetophenone. The best catalyst was an N^α-acylated dipeptide alkylamide with the -TOAC-(αMe)Val- sequence folded in a stable, intramolecularly hydrogen-bonded β-turn conformation with large, lipophilic (hydrophobic) N- and C-terminal blocking groups. We rationalized our findings by proposing models for the diastereomeric intermediates between (R)-[and (S)]-1-phenylethanol and the catalyst Fmoc-TOAC-L(αMe)Val-NHiPr, based on the X-ray diffraction structure of the latter.

Full text of DOI:10.1002/1521-3765(20020104)8:1<84::AID-CHEM84>3.0.CO;2-N

FULL PAPER
Nitroxyl Peptides as Catalysts of Enantioselective Oxidations
Fernando Formaggio,[a] Marcella Bonchio,[b] Marco Crisma,[a] Cristina Peggion,[a]
Stefano Mezzato,[a] Alessandra Polese,[a] Alessandra Barazza,[a] Sabrina Antonello,[c]
Flavio Maran,[c] Quirinus B. Broxterman,[d] Bernard Kaptein,[d] Johan Kamphuis,[e]
Rosa Maria Vitale,[f] Michele Saviano,[g] Ettore Benedetti,[g] and Claudio Toniolo*[a]
Abstract: The achiral, nitroxyl-contain-
ing a-amino acid TOAC (TOAC ˆ
2,2,6,6-tetramethylpiperidine-1-oxyl-4-
amino-4-carboxylic acid), in combina-
tion with the chiral a-amino acid Ca-
methyl valine [(aMe)Val], was used to
prepare short peptides (from di- to hexa-)
that induced the enantioselective oxi-
dation of racemic 1-phenylethanol to
acetophenone. The best catalyst was an
gen-bonded b-turn conformation with
large, lipophilic (hydrophobic) N- and
C-terminal blocking groups. We ration-
alized our findings by proposing models
for the diastereomeric intermediates
between (R)-[and (S)]-1-phenylethanol
and the catalyst Fmoc-TOAC-l-
(aMe)Val-NHiPr, based on the X-ray
diffraction structure of the latter.
Na-acylated dipeptide alkylamide with
À
À
the TOAC-(aMe)Val sequence fold-
ed in a stable, intramolecularly hydro-
Keywords: alcohols
¥ asymmetric
catalysis ¥ conformation analysis ¥
oxidation ¥ peptidomimetics
Ac6c (Ac6c ˆ 1-amino-1-cyclohexanecarboxylic acid), the cy-
Introduction
3]
cloaliphatic analogue of TOAC, strongly stabilize b-turn[1
TOAC (TOAC ˆ 2,2,6,6-tetramethylpiperidine-1-oxyl-4-ami-
no-4-carboxylic acid) is a member of the family of conforma-
tionally constrained Ca-tetrasubstituted a-amino acids, the
prototype of which is a-aminoisobutyric acid (Aib). Aib and
all the Cia $ Cia-cyclized a-amino acids of this family, like
and 310/a-helical[4] conformations in peptide molecules.[5, 6]
The structures of Aib, Ac6c and TOAC are given in Figure 1.
[a] Prof. C. Toniolo, Prof. F. Formaggio, Dr. M. Crisma, Dr. C. Peggion,
S. Mezzato, Dr. A. Polese, A. Barazza
Department of Organic Chemistry, University of Padova
CNR Centre CSB, via Marzolo 1, 35131 Padova (Italy)
Fax : (‡39)049-827-5239
E-mail: biop02@chor.unipd.it
[b] Dr. M. Bonchio
Department of Organic Chemistry, University of Padova
CNR Centre CMRO, 35131 Padova (Italy)
[c] Dr. S. Antonello, Prof. F. Maran
Department of Physical Chemistry, University of Padova
35131 Padova (Italy)
Figure 1. Chemical structures of the Ca-tetrasubstituted a-amino acids
Aib, l-(aMe)Val, Ac6c, and TOAC.
[d] Dr. Q. B. Broxterman, Dr. B. Kaptein
DSM Research, Organic Chemistry and Biotechnology Section
P.O. Box 18, 6160 MD Geleen (The Netherlands)
TOAC has a saturated heterocyclic structure containing a
paramagnetic probe (a nitroxyl group) stabilized by the
presence of two contiguous tetrasubstituted carbon atoms.[7]
A favourable property of TOAC, over other spin-labelled
amino acids connected to the peptide backbone through a
flexible link, is the very restricted mobility owing to the Ca-
tetrasubstitution and hampered rotation about side-chain
bonds; this is a result of the incorporation of the nitroxyl
nitrogen and the Ca, Cb and Cg atoms into a cyclic moiety.
Therefore, TOAC represents an extremely useful tool to
[e] Dr. J. Kamphuis
DSM Food Specialties, Nutritional Ingredients
P.O. Box 1, 2600 Delft (The Netherlands)
[f] Dr. R. M. Vitale
Department of Environmental Sciences
2nd University of Naples, 81100 Caserta (Italy)
[g] Dr. M. Saviano, Prof. E. Benedetti
Biocrystallography Research Centre, CNR
Department of Chemistry, University of Naples ™Federico II∫
80134 Naples (Italy)
84
¹ WILEY-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002
0947-6539/02/0801-0084 $ 17.50+.50/0
Chem. Eur. J. 2002, 8, No. 1

84 93
exploit sensitive electron spin resonance techniques for
peptide and protein studies.[8, 9] In addition, as a free radical,
TOAC is able to induce a dramatic intramolecular quenching
of suitably tailored, fluorescence labelled peptides.[10, 11] We
have also shown that TOAC undergoes a reversible, nitroxide-
based redox process that can be monitored by cyclic
voltammetry.[8]
crystal state by X-ray diffraction. The capability of these
peptides to act as mediators (catalysts) of chemical and
electrochemical enantioselective oxidations was tested on the
racemic secondary alcohol 1-phenylethanol. The results were
interpreted by performing conformational energy computa-
tions on the mechanistically critical reaction intermediate.
In the present work we take advantage of the redox
properties of TOAC-containing peptides by exploiting them
as potential catalysts for the enantioselective oxidation of a
secondary alcohol. As TOAC is an achiral amino acid residue,
we incorporated it into short peptides based on the chiral Ca-
tetrasubstituted (aMe)Val (Ca-methylvaline) (Figure 1). We
chose this chiral residue because: 1) in analogy with Aib, Ac6c
Results and Discussion
Synthesis and characterization: The synthesis of TOAC is
described in references [7, 8]. For the large-scale production
of enantiomerically pure l-(aMe)Val, an economically at-
tractive and generally applicable chemo-enzymatic synthesis,
developed by DSM Research, was exploited.[18, 19] This
involved a combination of a partial Strecker synthesis, for
the preparation of the racemic a-amino amide, followed by
the use of the broadly specific a-amino amidase to achieve
optical resolution. All peptides were prepared by solution
methods. The homo-peptide series Z-[l-(aMe)Val]n-NHtBu
(n ˆ 2 4) was synthesized by using the acyl fluoride C-acti-
vation method,[20] which gives good yields for the coupling of
two sterically demanding (aMe)Val residues. Z-l-(aMe)Val-
F[17] was prepared from the N-protected amino acid[13, 16] and
cyanuric fluoride in pyridine. The various Z-l-(or d-)(aMe)-
Val-NHR (R ˆ Me, Et, iPr, tBu) derivatives were synthesized
by treating the Na-protected amino acid with the relevant
primary amine in the presence of EDC (EDC ˆ 1-(3-dimeth-
7, 12 17]
and TOAC, it is a strong b-turn/310-helical inducer,[5
2) it contains an intrinsic source of chirality (the asymmetric
a-carbon) and 3) among the chiral Ca-tetrasubstituted a-
amino acids, it has the strongest known bias towards a specific
turn/helix screw sense (right-handed for the l-enantiomer)
17]
(peptide overall molecular dissymmetry).[15
We synthesized, by solution methods, a set of linear TOAC/
l-(or d-)(aMe)Val-containing di- to hexapeptides (1 11 and
13 16) with different Na-protecting/blocking groups [Fmoc
(Fmoc ˆ fluoren-9-ylmethoxycarbonyl),
Z
(Z ˆ benzyloxy-
carbonyl), Boc (Boc ˆ tert-butoxycarbonyl), or Ac (Ac ˆ
acetyl)] and C-blocking/protecting groups (NHtBu, NHiPr,
NHEt, NHMe, or OMe), in which the TOAC residue was
inserted either at the N- or C-terminus, or in an internal
position of the peptide sequence. The reference dipeptides
Fmoc-TOAC-l-Val-NHtBu (12) and cyclo[TOAC-l-(aMe)-
Val] (17) were also prepared. The preferred conformations of
these compounds in CDCl3 were examined by IR spectros-
copy and, for dipeptide 1 and pentapeptide 15, also in the
ylamino)propyl-3-ethylcarbodiimide).
Benzyloxycarbonyl
Na-deprotection was carried out by catalytic hydrogenolysis.
The TOAC Fmoc,[21] Boc,[22] and Z[23] Na derivatives were
obtained according to published procedures. All coupling
steps involving TOAC were performed by exploiting the
highly efficient EDC/HOAt (HOAt ˆ 1-hydroxy-7-aza-1,2,3-
benzotriazole) C-activation method.[24] Fmoc Na-deprotection
was achieved by treatment with a diethylamine solution
(25% v/v). The Na-acetylated peptide was obtained by
coupling the Na-deprotected synthetic precursor with acetic
anhydride. The Na-free dipeptide ester H-TOAC-l-(aMe)-
Val-OMe was cyclized to the 2,5-dioxopiperazine cyclo-
[TOAC-l-(aMe)Val] (17) by heating at reflux in a toluene/
acetic acid 20:1 (v/v) mixture. The synthesis and character-
ization of the penta- and hexapeptides Fmoc-[l-(aMe)Val]n-
TOAC-[l-(aMe)Val]2-NHtBu (15: n ˆ 2; 16: n ˆ 3) are al-
ready published.[11]
Claudio Toniolo[*] received his Laurea
in chemistry from the University of
Padova, Italy, in 1965. Following post-
doctoral study with Prof. M. Goodman
at the Department of Chemistry, Sec-
tion of Biopolymers, Polytechnic Insti-
tute of Brooklyn, New York, 1967-8, he
joined the Faculty of Sciences, Univer-
sity of Padova, where he has been
Professor of Organic Chemistry since
All peptides were characterized by melting point and
optical rotatory power determinations, thin-layer chromatog-
raphy (TLC) in three solvent systems and solid-state IR
absorption (Table 1).
1980. He has been a Visiting Scholar or Professor at Ports-
mouth Polytechnic, Portsmouth, UK (Prof. E. M. Bradbury),
the State University of New York at Binghamton (Prof. E. S.
Stevens), the Indian Institute of Sciences, Bangalore (Prof. P.
Balaram), University of California at San Diego (Prof. M.
Goodman), Osaka University (Prof. Y. Kobayashi) and the
National University of Somalia, Mogadishu. He is a member of
the editorial board of several peptide-science and bioorganic-
chemistry journals and co-author of over 500 publications on
peptide synthesis, supramolecular chemistry and three-dimen-
sional structure.
Conformational analysis: The preferred conformation adopt-
ed by the linear, Na-acylated TOAC/(aMe)Val dipeptide
amides, in a structure supporting solvent (CDCl3), was
1
investigated by FTIR spectroscopy. H NMR spectroscopy
can not routinely be used with TOAC peptides because the
nitroxyl group broadens the resonances of nearby protons
beyond detection (radical-induced relaxation effect). Figure 2
À
illustrates the FTIR absorption spectra in the N H stretching
[*] Members of the Editorial Board will be introduced to readers with their
first manuscript.
region of three relevant Na-acylated dipeptide amides. The
Chem. Eur. J. 2002, 8, No. 1
¹ WILEY-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002
0947-6539/02/0801-0085 $ 17.50+.50/0
85

C. Toniolo et al.
FULL PAPER
Table 1. Physical properties and analytical data for new TOAC and (aMe)Val derivatives and peptides.
[f]
Compound
M.p.[8C][a] Recryst.
[a]2D0[c]
TLC[e]
Rf(I) Rf(II) Rf(III)
IR u [cmÀ1
]
solvent[b]
Fmoc-TOAC-NHtBu
Z-l-(aMe)Val-NHtBu
Z-[l-(aMe)Val]2-NHtBu
Z-[l-(aMe)Val]3-NHtBu
Z-[l-(aMe)Val]4-NHtBu
Z-d-(aMe)Val-NHtBu
Z-l-(aMe)Val-NHiPr
Z-l-(aMe)Val-NHEt
138 140
116 117
126 127
162 163
245 246
115 117
92 94
EtOAc/PE
EtOAc/PE
EtOAc/PE
EtOAc/PE
DCM/PE
EtOAc/PE
EtOAc/PE
EtOAc/PE
EtOAc/PE
EtOAc/PE
-
8.1
12.2
18.4
17.8
À 8.0
0.95 0.95
0.95 0.95
0.95 0.95
0.95 0.95
0.95 0.90
0.95 0.95
0.45
0.50
0.40
0.35
0.30
0.50
0.50
0.40
0.40
0.40
0.40
0.40
0.35
0.40
0.35
0.35
0.25
0.20
0.20
0.30
0.40
0.50
0.40
0.40
3334, 1727, 1702, 1678, 1523
3417, 3298, 1722, 1663, 1538
3359, 3296, 1713, 1685, 1657, 1522
3343, 3309, 1702, 1673, 1654, 1515
3340, 1707, 1668, 1528
3418, 3299, 1718, 1659, 1535
3383, 3327, 3297, 1727, 1708, 1693, 1660, 1645, 1539
3344, 3316, 1708, 1635, 1535
3334, 1708, 1653, 1526
3341, 1725, 1652, 1522
3340, 1722, 1652, 1520
3371, 1702, 1631, 1527
3378, 1701, 1636, 1539
3427, 3319, 1718, 1664, 1523
3434, 3378, 3289, 1710, 1684, 1658, 1521
3394, 1707, 1681, 1637, 1523
3338, 1694, 1669, 1658, 1510
3351, 3337, 1691, 1671, 1659, 1646, 1532
3343, 1664, 1523
3441, 3383, 1677, 1660, 1533
3341, 1725, 1652, 1522
3323, 1743, 1721, 1660
À 0.4[d] 0.95 0.95
0.4[d] 0.90 0.90
À 0.6[d] 0.80 0.90
oil
oil
Z-l-(aMe)Val-NHMe
Fmoc-TOAC-l-(aMe)Val-NHtBu (1)
Fmoc-TOAC-d-(aMe)Val-NHtBu (2)
Fmoc-TOAC-l-(aMe)Val-NHiPr (3)
Fmoc-TOAC-l-(aMe)Val-NHEt (4)
Fmoc-TOAC-l-(aMe)Val-NHMe (5)
Z-TOAC-l-(aMe)Val-NHtBu (6)
Boc-TOAC-l-(aMe)Val-NHtBu (7)
Boc-TOAC-[l-(aMe)Val]2-NHtBu (8)
Boc-TOAC-[l-(aMe)Val]3-NHtBu (9)
165 167
165 167
155 157
162 164
158 160
156 157
208 209
213 214
251 252
14.8
0.90 0.95
0.90 0.95
0.75 0.95
0.75 0.95
0.70 0.90
0.75 0.95
0.80 0.95
0.90 0.95
0.95 0.90
0.95 0.85
0.65 0.85
0.90 0.95
EtOAc/PE À 14.9
EtOAc/PE
EtOAc/PE
EtOAc/PE
EtOAc/PE
EtOAc/PE
DCM/PE
18.8
19.7
23.5
41.2
40.8
58.6
55.2
53.2
46.4
14.6
DCM/PE
DCM/PE
Boc-TOAC-[l-(aMe)Val]4-NHtBu (10) 244 245
Ac-TOAC-l-(aMe)Val-NHtBu (11)
Fmoc-TOAC-l-Val-NHtBu (12)
Fmoc-TOAC-l-(aMe)Val-OMe (13)
Fmoc-l-(aMe)Val-TOAC-NHtBu (14) 131 133
cyclo[TOAC-l-(aMe)Val] (17) 240 242
181 182
165 167
165 167
EtOAc/PE
EtOAc/PE
EtOAc/PE
EtOAc/PE
hot toluene
1.3[d] 0.95 0.95
4.3[d] 0.95 0.95
À 8.9[d] 0.80 0.85
3365, 1720, 1670, 1525
3426, 3315, 1662, 1557
[a] Determined on a Leitz model Laborlux 12 apparatus (Wetzlar, Germany). [b] EtOAc ˆ ethyl acetate, PE ˆ petroleum ether, DCM ˆ dichloromethane.
[c] Determined on a Perkin Elmer model 241 polarimeter (Norwalk, CT) equipped with a Haake model L thermostat (Karlsruhe, Germany): c ˆ 0.5
20
(methanol). [d] [a] . [e] Silica gel plates (60F-254 Merck), solvent systems: I) chloroform/ethanol 9:1, II) butan-1-ol/water/acetic acid 3:1:1, III) toluene/
546
ethanol 7:1; the plates were developed with a UV lamp or with the hypochlorite/starch/iodide chromatic reaction, as appropriate; a single spot was observed
in each case. [f] Determined in KBr pellets on a Perkin Elmer model 580B spectrophotometer equipped with a Perkin Elmer model 3600 IR data station.
curves are characterized by two bands, at around 3435 cmÀ1
(free, solvated NH groups) and 3380 cmÀ1 (H-bonded NH
groups).[25, 26] The spectra do not vary appreciably with
changing peptide concentration (in the range 10 0.1mm)
(not shown). Therefore, the observed hydrogen bonding
ˆ
À
arises almost exclusively from intramolecular C O ¥¥¥ H N
interactions. The remarkable intensity of the low-frequency
band shown by all the Na-acylated TOAC/(aMe)Val dipeptide
amides (for peptide 1 see Figure 2, curve A) suggests a
significant population of intramolecularly H-bonded species
in these compounds. Interestingly, this band is: 1) much less
Abstract in Italian: L×a-amminoacido achirale TOAC, conte-
nente un gruppo nitrossilico, in combinazione con l×a-ammi-
noacido chirale Ca-metil-valina [(aMe)Val] õ stato utilizzato
per sintetizzare peptidi corti (dal di- all×esapeptide) capaci di
indurre ossidazione enantioselettiva del racemato dell×1-feni-
letanolo ad acetofenone. Il miglior catalizzatore õ risultato
essere un dipeptide alchilammide Na-acilato e caratterizzato
À
Figure 2. FTIR absorption spectra in the N H stretching region (3500
3350 cmÀ1 region) of A) Fmoc-TOAC-l-(aMe)Val-NHtBu (1), B) Fmoc-
TOAC-l-Val-NHtBu (12), and C) Fmoc-l-(aMe)Val-TOAC-NHtBu (14)
in CDCl3 solution (conc. 1mm).
strong in the TOAC/Val analogue 12 (see curve B), high-
lighting the positive effect of Ca-tetrasubstitution [cf.(aMe)-
Val vs Val] on intramolecular H-bond formation [6, 26] and 2) of
appreciable intensity in the (aMe)Val/TOAC dipeptide 14
(see curve C) indicating that a set of two consecutive Ca-
tetrasubstituted a-amino acids, but not necessarily their
precise sequence, is important for a high content of intra-
À
À
dalla sequenza TOAC-(aMe)Val , che assume una confor-
mazione ripiegata-b stabilizzata da un legame a idrogeno
ˆ
À
intramolecolare C O ¥¥¥ H N, e da gruppi bloccanti N- e
C-terminali largamente lipofilici (idrofobici). Questi risultati
sono stati razionalizzati sulla base dei modelli degli intermedi
diastereomerici tra l×(R) [e l×(S)]-1-feniletanolo e il catalizza-
tore Fmoc-TOAC-l-(aMe)Val-NHiPr, elaborati partendo dal-
la struttura tridimensionale del dipeptide ottenuta dall×analisi
della diffrazione dei raggi X.
molecular H-bonding. The known conformational preferen-
17]
ces of TOAC[7] and (aMe)Val,[12
combined with the
position of the low-frequency band, strongly support the view
that the intramolecularly H-bonded species formed by the Na-
86
¹ WILEY-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002
0947-6539/02/0801-0086 $ 17.50+.50/0
Chem. Eur. J. 2002, 8, No. 1

Nitroxyl Peptides as Catalysts
84 93
Table 2. Relevant backbone torsion angles [8] for Fmoc-TOAC-l-(aMe)-
Val-NHtBu (1) and Fmoc-[l-(aMe)Val]2-TOAC-[l-(aMe)Val]2-NHtBu
(15) with esds in parentheses.
acylated TOAC/(aMe)Val dipeptide amides 1 7 and 11 are
of the b-turn type. It is reasonable to assume that the
conformation of the longer TOAC/(aMe)Val peptides 8 10
(from tri- to hexapeptides) would evolve into two consecutive
b-turns and eventually to the 310-helix. Indeed, this latter
ordered secondary structure has characterized the folding in
solution of the penta- and hexapeptides 15 and 16, respec-
tively.[11]
Torsion angle
Dipeptide 1
Pentapeptide 15
q2
q1
À 116.7(3)
À 175.8(2)
À 178.4(2)
À 59.5(3)
À 31.7(3)
À 173.6(2)
À 57.6(3)
À 32.4(4)
À 175.5(3)
À 152.8(3)
178.7(2)
w0
f1
y1
w1
f2
y2
w2
f3
y3
w3
f4
y4
w4
f5
y5
w5
À 157.9(2)
À 62.8(3)
À 32.9(3)
À 173.1(2)
À 54.8(3)
À 28.2(3)
À 174.2(2)
À 50.5(3)
À 36.4(3)
À 168.8(2)
À 55.9(3)
À 29.9(3)
À 175.0(2)
À 61.8(3)
À -31.7(4)
164.9(4)
In addition, we used X-ray diffraction to determine the
preferred crystal-state conformation of the dipeptide amide
Fmoc-TOAC-l-(aMe)Val-NHtBu (1) and the pentapeptide
amide
Fmoc-[l-(aMe)Val]2-TOAC-[l-(aMe)Val]2-NHtBu
(15). These molecular structures are given in Figures 3 and
4, and relevant backbone torsion angles are given in Table 2.
Table 3 lists the intra- and intermolecular H-bond parameters.
Bond lengths and bond angles (deposited) are in good
agreement with previously reported values for Fmoc-ure-
thane[27] and NHtBu amide groups,[28] the peptide unit,[29, 30]
10, 31, 32]
and the TOAC[7
particular, the N
and (aMe)Val[12, 15, 16] residues. In
d
d
À
O
bond length of the TOAC nitroxyl
group (1.277(3) ä for 1 and 1.278(3) ä for 15), the external
Od-Nd-Cg bond angles (in the range 116.0(2) 119.7(2)8), and
the internal bond angle at the Nd atom (124.6(2)8 for 1 and
123.1(2)8 for 15) compare well with those published for other
TOAC residues.
All l-(aMe)Val and TOAC residues of the two peptides are
found in the right-handed helical region of the conformational
map. The average f and y values are À578 and À328,
respectively,[33] and are quite close to those typical for a right-
handed 310-helix (À57, À308).[4] Dipeptide amide 1 forms a
regular type-III b-turn.[1, 3] The single 1 4 intramolecular
ˆ
À
C O ¥¥¥ H N hydrogen bond has the NT O0 distance at the
36]
upper limit of the expected range (2.8 3.2 ä).[34
The
pentapeptide amide 15 is folded in a right-handed 310-helical
structure stabilized by four consecutive 1 4 intramolecular
ˆ
À
C O ¥¥¥ H N hydrogen bonds (the N O distances are in the
range 3.049(4) 3.235(3) ä).
All urethane, peptide and amide groups of the two
compounds are in the trans disposition. Interestingly, in
peptide 15 the N-terminal urethane w0, the peptidew3 and the
C-terminal amidew5 torsion angles deviate significantly
(jDw j> 118) from the trans pla-
Figure 3. X-ray diffraction structure of Fmoc-TOAC-l-(aMe)Val-NHtBu
ˆ
À
(1). The intramolecular C O ¥¥¥ H N hydrogen bond is represented by a
dashed line.
narity (1808). In both compounds
the Fmoc-urethane group (q1 and
w0 torsion angles) is in the usual
trans,trans conformation.[27]
Relevant
torsion
TOAC
of the two peptides
angles
piperidine
for
ring[7
the
10, 31, 32]
are listed in Table 4. Two sets of
values are observed for the tor-
sion angles relating the piperidine
ring to the peptide chain N-Ca-Cb-
Cg (c1) and C'-Ca-Cb-Cg. In the
conformation 608, À 608/1808,1808
adopted by dipeptide 1, the a-
amino group occupies the axial
position and the a-carboxyl group
occupies the equatorial position.
Figure 4. X-ray diffraction structure of Fmoc-[l-(aMe)Val]2-TOAC-[l-(aMe)Val]2-NHtBu (15). The intra-
ˆ
À
molecular C O ¥¥¥ H N hydrogen bonds are represented by dashed lines.
Chem. Eur. J. 2002, 8, No. 1
¹ WILEY-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002
0947-6539/02/0801-0087 $ 17.50+.50/0
87

C. Toniolo et al.
FULL PAPER
Table 3. Intra- and intermolecular hydrogen bond parameters for Fmoc-TOAC-l-(aMe)Val-NHtBu (1) and Fmoc-[l-(aMe)Val]2-TOAC-[l-(aMe)Val]2-
NHtBu (15).
Type of H-bond
Donor (D)
Acceptor (A)
Symmetry
operation
Distance [ä]
D ¥¥¥ A
Distance [ä]
H ¥¥¥ A
Angle [8]
À
D H ¥¥¥ A
À
1
intramolecular
intermolecular
intramolecular
NT H
À
N1 H
O0
O2
O0
O1
O2
O3
O3D
x, y, z
3.255(3)
3.205(3)
3.151(4)
3.235(3)
3.163(3)
3.049(4)
2.976(4)
2.459(2)
2.610(2)
2.305(4)
2.380(3)
2.329(3)
2.214(4)
2.136(4)
154.1(2)
127.3(2)
168.0(3)
173.0(3)
163.7(3)
163.9(3)
165.3(3)
1
3
x À ³2, ³2 À y, 2 À z
À
15
N3 H
x, y, z
À
N4 H
x, y, z
x, y, z
x, y, z
1 À x, y À ³2, 1 À z
À
N5 H
À
NT H
1
À
intermolecular
N1 H
Table 4. Relevant torsion angles [8] for the piperidine ring of the TOAC
residue in Fmoc-TOAC-l-(aMe)Val-NHtBu (1) and Fmoc-[l-(aMe)Val]2-
TOAC-[l-(aMe)Val]2-NHtBu (15).
Enantioselective chemical oxidation of a secondary alcohol:
Efficient methods for the chemical oxidation of alcohols
40]
mediated by a nitroxyl derivative have been reported.[38
Torsion angle
Dipeptide 1
Pentapeptide 15
The one-electron oxidation of a nitroxyl compound (I) by
OBrÀ (arising from the reaction of bleach with a bromide ion)
affords an N-oxoammonium cation (II) to which a secondary
alcohol, such as 1-phenylethanol (III), can add (Scheme 1).
The resulting intermediate IV undergoes elimination through
a cyclic dipolar mechanism, thereby producing the ketone V
and the hydroxylamine derivative VI. Re-oxidation of VI to II
completes the catalytic cycle. Enantioselective chemical
oxidations of racemic secondary alcohols, using optically
N-Ca-Cb-Cg (c1)
À 73.6(3)
69.7(3)
À 145.3(2)
87.0(3)
C'-Ca-Cb-Cg
167.6(2)
À 167.6(2)
À 162.1(3)
160.9(3)
51.0(3)
À 52.4(3)
À 43.6(3)
45.8(3)
96.1(3)
À 151.2(3)
160.8(3)
148.1(3)
À 24.1(3)
À 31.9(3)
51.3(3)
Od-Nd-Cg-Cb
Cg-Cb-Ca-Cb' (q1, q1')
Ca-Cb-Cg-Nd (q2, q2')
Cb-Cg-Nd-Cg' (q3, q3')
57.3(3)
active nitroxyl radicals as catalysts, have recently been
39.0(4)
À 40.1(4)
À 23.6(4)
43]
À 27.4(4)
performed.[41
However, in general, the synthesis of the
chiral nitroxyl compound is very laborious and some of these
catalysts undergo facile racemization.[44]
In the other conformation 908, À 1508/ À 1508,908, exhibited
by pentapeptide 15, the a-amino and a-carboxyl substituents
are located at intermediate positions between axial and
equatorial, the latter being characterized by the torsion angles
1808,1808/ À 608,608. Except for the q1, q1', q3 and q3' torsion
angles of pentapeptide 15, all other endocyclic torsion angles
for the two peptides show values in the Æ39 588 range, close
to those of piperidine and cyclohexane. These findings
indicate that the piperidine ring of pentapeptide 15 is more
flattened than that of dipeptide 1 in both the Ca and Nd
regions. The piperidine ring of dipeptide 1 adopts an
approximate chair conformation with the following puckering
parameters:[37] QT ˆ 0.46(1) ä, f2 ˆ 172(2)8 and q2 ˆ 13(1)8,
while that of pentapeptide 15 adopts an approximate twist-
boat conformation, with one Cb atom above and the other Cb
atom below the average plane of the ring, with the following
puckering parameters: QT ˆ 0.66(1) ä, f2 ˆ 91(1)8 and q2 ˆ
d
d
g
d
g'
À
92(1)8. The angle between the N O bond and the C -N -C
plane is 18.8(3)8 for dipeptide 1 and 3.9(3)8 for pentapeptide
15. Therefore, the C(C)-N-O group adopts a pyramidal shape
in the chair form of dipeptide 1 only. The angles between the
normals to the average planes of the Fmoc and TOAC rings
are 35.4(1)8 in dipeptide 1 and 56.8(1)8 in pentapeptide 15.
In the crystals of dipeptide 1, molecules pack in the unit cell
in rows parallel to the a direction through weak (urethane)
Scheme 1. Catalytic cycle for the oxidation of a secondary alcohol using a
bulk oxidant and a nitroxyl catalyst.
À
ˆ
À
N1 H ¥¥¥ O2 C2 (amide) hydrogen bonds. The peptide N2 H
moiety does not seem to be involved in the hydrogen bonding
array. Along the b direction, the helical molecules of
pentapeptide 15 are connected by hydrogen bonds between
The results of the enantioselective chemical oxidation of
racemic 1-phenylethanol to acetophenone, by means of the
chiral, non-racemizable TOAC/(aMe)Val peptides as cata-
lysts, are listed in Table 5. As expected, the enantiomeric
dipeptide catalysts 1 and 2 give the same enantiomeric excess
of recovered alcohol [the (R)-alcohol enantiomer is oxidized
À
À
the urethane N1 H donor and the nitroxyl O3D N3D
acceptor groups.
88
¹ WILEY-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002
0947-6539/02/0801-0088 $ 17.50+.50/0
Chem. Eur. J. 2002, 8, No. 1

Nitroxyl Peptides as Catalysts
84 93
Table 5. Enantioselective chemical oxidation of racemic 1-phenylethanol with the TOAC/(aMe)Val peptide catalysts.
Catalyst
ee [%]
Configuration[a]
Conversion (C) [ %]
S[b]
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
Fmoc-TOAC-l-(aMe)Val-NHtBu
Fmoc-TOAC-d-(aMe)Val-NHtBu
Fmoc-TOAC-l-(aMe)Val-NHiPr
Fmoc-TOAC-l-(aMe)Val-NHEt
Fmoc-TOAC-l-(aMe)Val-NHMe
Z-TOAC-l-(aMe)Val-NHtBu
Boc-TOAC-l-(aMe)Val-NHtBu
Boc-TOAC-[l-(aMe)Val]2-NHtBu
Boc-TOAC-[l-(aMe)Val]3-NHtBu
Boc-TOAC-[l-(aMe)Val]4-NHtBu
Ac-TOAC-l-(aMe)Val-NHtBu
67
67
69
52
50
75
70
62
74
68
60
33
13
10
36
38
18
S(À)
R(‡)
S(À)
S(À)
S(À)
S(À)
S(À)
83
82
82
83
84
82
82
79
81
81
85
81
82
83
80
79
81
2.3
2.3
2.4
1.9
1.8
2.7
2.5
2.1
2.7
2.4
2.0
1.5
1.2
1.1
1.6
1.6
1.2
À
S( )
À
S( )
S(À)
S(À)
S(À)
R(‡)
R(‡)
S(À)
R(‡)
S(À)
Fmoc-TOAC-l-Val-NHtBu
Fmoc-TOAC-l-(aMe)Val-OMe
Fmoc-l-(aMe)Val-TOAC-NHtBu
Fmoc-[l-(aMe)Val]2-TOAC-[l-(aMe)Val]2-NHtBu
Fmoc-[l-(aMe)Val]3-TOAC-[l-(aMe)Val]2-NHtBu
cyclo[TOAC-l-(aMe)Val]
[a] Of recovered alcohol. [b] S ˆ ln[(1 À C)(1 À ee)]/ln[(1 À C)(1‡ee)].
preferentially by the l-(aMe)Val peptide]. Therefore, it is
possible to get a mixture enriched in either one of the two
enantiomeric alcohols by using the catalyst of the appropriate
chirality. The highest enantiomeric excesses (ee) and selec-
tivity factors (S) were obtained when both N- and C-blocking
groups were highly lipophilic (hydrophobic) (compare the
catalyst series 1, 3 5 and 6, 7, 11). Replacement of the turn-
inducer (aMe)Val (in catalyst 1) with Val, which is more
prone to adopt partially extended conformations, results in a
dramatic decrease in the efficiency of the kinetic resolution
(see catalyst 12). An analogous result is obtained when the
alkylamide moiety at the C-terminus (of catalyst 1) is replaced
by an ester functionality, which precludes intramolecularly
H-bonded b-turn formation (see catalyst 13). Interestingly, in
this case the final mixture was enriched in the (R)-alcohol and
not in the (S)-alcohol as was found for catalysts 1 and 3 12.
Also, the b-turn-forming (aMe)Val/TOAC sequence permu-
tation (catalyst 14) produced a low R enantiomeric excess.
Lengthening the main-chain of the peptide with two, three
and four l-(aMe)Val residues at the C-terminus (catalysts 8
10), does not improve enantioselectivity with respect to the
parent dipeptide 7. Modest enantioselectivities were observed
for the long penta- and hexapeptides with an internal TOAC
residue (catalysts 15 and 16). The TOAC-based cyclic dipep-
tide 17 was a poor catalyst.
electro-oxidations.[49] Such a procedure, in which electrolysis
is carried out in an undivided cell under constant current
conditions, combines the advantages of practical simplicity
and low environmental stress. Indeed, in this case the use of
highly concentrated supporting electrolyte solutions (quater-
nary ammonium salts), which are required to carry out the
process in divided cells, is unnecessary. Very recently, a further
improvement has been described in which the organic solvent
is replaced by an aqueous silica-gel disperse system.[50]
The indirect electro-oxidation in an undivided cell is based
on a CH2Cl2/H2O two-phase system employing two mediators.
A systematic investigation of the different factors that affect
the efficiency of the oxidation under these conditions has
been carried out.[51] The reaction conditions have been
optimized by using a concentration of NaBr in the aqueous
solution >15w%, a pH of 8.6 (saturated NaHCO3 buffer
solution), a concentration of the nitroxyl mediator in the
organic phase ꢀ1 mol% with respect to the alcohol and a
current density of 10 140 mAcmÀ2 with a Pt electrode. It has
been observed, however, that the current efficiency is not
unitary and that oxidations are essentially completed after
consumption of 2.2 2.6 FmolÀ1 of charge.
In our electrochemical experiments we used a 20 w% NaBr
aqueous solution saturated with NaHCO3, a nitroxyl/alcohol
ratio of about 0.017 and a current density of 10 mAcmÀ2.
Bromide ions in the aqueous solution undergo electro-
oxidation at the Pt anode and subsequently oxidize the
nitroxyl in the organic phase; electron transfer takes place at
the CH2Cl2/H2O interface. The active form of the mediator
can then oxidize the racemic alcohol. The use of a two-phase
electrochemical system allowed us to carry out the electro-
chemical experiments under conditions that better resembled
those described above for the chemical oxidations.
The main conclusion drawn from this investigation is that
the best peptide catalysts investigated have large lipophilic
(hydrophobic) blocking groups at either terminus, an N-ter-
minal TOAC residue and are folded in a stable, intramolec-
ˆ
À
ularly C O ¥¥¥ H N hydrogen-bonded b-turn conformation.
Enantioselective electrochemical oxidation of a secondary
alcohol: Some results on the electrochemical kinetic resolu-
tion of racemic secondary alcohols have been reported, either
by use of synthetically complex, optically active nitroxyl
mediators or nitroxyl-modified electrodes.[45 48] Aprotic media
and divided cells are usually employed in these experiments,
probably because the N-oxoammonium salts are labile in
water. Only recently, however, an aqueous/organic, two-phase
system has efficiently been exploited in enantioselective
The catalytic behaviour of the Boc-TOAC-l-(aMe)Val-
NHtBu mediator was analyzed at 08C by measuring the ee of
the alcoholic mixture at different degrees of conversion (C).
The results (Table 6) display some scatter of the selectivity
factor (S) values. However, a reasonably good plot of ee
against C was obtained leading to a calculated average S value
of 1.8. As reported by Tanaka and co-workers,[49] an increase
Chem. Eur. J. 2002, 8, No. 1
¹ WILEY-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002
0947-6539/02/0801-0089 $ 17.50+.50/0
89

C. Toniolo et al.
FULL PAPER
Table 6. Enantioselective electrochemical oxidation of racemic 1-phenyl-
ethanol with the TOAC/(aMe)Val peptide catalysts.
ethanol and Fmoc-TOAC-l-(aMe)Val-NHtBu. Models were
constructed for the diastereomers considering the two possi-
ble directions of addition of alcohol (III) to the TOAC N-
oxoammonium double bond of (II). The maps show that the
two diastereomeric intermediates IV, with addition of the
alcohol to the TOAC ring on the side away from the Fmoc
group, exhibit the same energy (within experimental error). In
contrast, the two diastereomeric intermediates (IV) from
addition to the ring side close to the Fmoc group are more
stable than those discussed above, and the intermediate from
(R)-1-phenylethanol presents a conformation with an energy
4.4 KcalmolÀ1 lower than that computed for the other
diastereomer. These findings are confirmed by the mini-
mum-energy conformations obtained after a full minimization
procedure of the low-energy conformations for the two
models. Figure 5 illustrates the stereoviews for the minimum
energy structures of the ™close∫ diastereomeric intermediates.
Our computational analysis indicates that the phenyl group of
(R)-1-phenylethanol (Figure 5B) is involved in a favourable,
intramolecular, edge-to-face, aromatic aromatic interaction
with the Fmoc moiety of the dipeptide catalyst (the angle
between the normals to the planes of the phenyl and fluorenyl
systems is 1158).[52] We believe this type of interaction is
important for high diastereoselectivity. Interestingly, this
finding is in excellent agreement with the conclusions recently
put forward to explain the unexpected diastereoselectivity of
the Sharpless asymmetric dihydroxylation of allyl d-xylo-
sides,[53] which highlight the essential role of aromatic or, more
Catalyst
ee
[%] tion[a]
Configura- Conversion S[b]
(C)[%]
2
7
Fmoc-TOAC-d-(aMe)Val-NHtBu 25
R(‡)
S(À)
S(À)
S(À)
S(À)
R(‡)
66
53
59
76
84
55
1.6
1.5
1.8
2.1
1.8
1.1
Boc-TOAC-l-(aMe)Val-NHtBu
15
25
52
53
2
13 Fmoc-TOAC-l-(aMe)Val-OMe
[a] Of recovered alcohol. [b] S ˆ ln[(1 À C)(1 À ee)]/ln[(1 À C)(1‡ee)].
of selectivity can be induced by decreasing the temperature.
Indeed, we obtained an S value of 3.1 at À108C. The trend of
selectivity for the three mediators tested resembles that
resulting from the corresponding chemical oxidations, but in
general a lower efficiency of the alcohol resolution was
observed under the electrochemical conditions.
Conformational energy calculations: To understand the
factors playing a role in the stereoselectivity of 1-phenyl-
ethanol oxidation catalyzed by chiral peptides that contain
TOAC residues, we performed energy minimization calcula-
tions. As a first step, we investigated the conformational
behaviour of Fmoc-TOAC-l-(aMe)Val-NHtBu (1) and com-
pared the results with those of the simple dipeptide reference
compound Ac-l-Ala-l-Ala-NHtBu. Our data indicated that
the minimized Fmoc-TOAC- l-(aMe)Val-NHtBu (1) back-
bone structure (f1 ˆ À62.18,
y1 ˆ À14.18; f2 ˆ À45.88, y2 ˆ
À38.58) is characterized by a
type-III b-turn conformation,
closely resembling the crystal-
state structure described above,
with a root-mean-square devia-
tion for all heavy atoms of
0.53 ä. In contrast, the analysis
of the minimized structure of
the Ac-l-Ala-l-Ala-NHtBu re-
vealed the absence of the b-turn
conformation which was instead
replaced by a more stable (DE ˆ
1.5 KcalmolÀ1) structure with
two consecutive g-turns (f1 ˆ
69.98, y1 ˆ À63.98; f2 ˆ À83.88,
y2 ˆ 82.88).[2] These findings
agree with the known strong
preference of the two sterically
demanding, Ca-tetrasubstituted
a-amino acids in the Fmoc-
TOAC-l-(aMe)Val-NHtBu di-
peptide for the b-turn conforma-
tion.
Subsequently, we obtained
rigid rotor maps for the torsion
À
angles around the N O and
À
O C bonds of the diastereomer-
Figure 5. Stereoviews of the minimum energy structures of the diastereomeric intermediates IV (Scheme 1)
between Fmoc-TOAC-l-(aMe)Val-NHtBu (1) and A) (S)-1-phenylethanol and B) (R)-1-phenylethanol, with
addition of the alcohol on the TOAC ring side close to the Fmoc group.
ic intermediates IV (Scheme 1)
between (R) [and (S)]-1-phenyl-
90
¹ WILEY-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002
0947-6539/02/0801-0090 $ 17.50+.50/0
Chem. Eur. J. 2002, 8, No. 1

Nitroxyl Peptides as Catalysts
84 93
Table 7. Crystallographic data and structure refinement for Fmoc-TOAC-l-
(aMe)Val-NHtBu (1) and Fmoc-[l-(aMe)Val]2-TOAC-[l-(aMe)Val]2-NHtBu
(15).
generally, large lipophilic (hydrophobic) moieties of the
substrate and catalyst in interactions leading to preferential
stabilization of one diastereomeric intermediate.
Peptide 1
Peptide 15
formula
Mr
T [K]
l [ä]
C35H49N4O5
605.8
293(2)
C53H82N7O8
945.3
293(2)
Conclusion
1.5418
1.5418
Considerable recent interest has been focused on the enan-
tioselective oxidation of racemic alcohols with chiral nitroxyl
catalysts.[41 51] In this paper we have shown that TOAC-based,
conformationally rigid, stable b-turn forming, very short
peptides are reasonably valuable catalysts in chemical oxida-
tions and, although less efficient, in electrochemical oxida-
tions as well. Longer helical structures are not required. Large
aromatic or aliphatic N- and C-blocking groups and the
N-terminal positioning of TOAC have a positive effect on the
efficiency. We have explained these experimental findings on
the basis of computed models for the intermediate adducts
between the chiral dipeptide amide N-oxoammonium medi-
ator and the enantiomeric secondary alcohol substrates. The
identity of the fast reacting alcohol enantiomer has been
correctly predicted.
However, it is clear that the experimental selectivity factors
(in the range 2.3 2.7 for our best catalysts, which corresponds
to a free-energy difference of only about 0.5 kcalmolÀ1),
although encouraging for a first-generation peptide system,
are far from optimal, particularly in view of the high
conversion (81 83%) considered in our enantioselective
chemical oxidation experiments. It is our contention that
these results are related to the only chiral centre of the
catalyst, that is, the (aMe)Val a-carbon atom, being too far
removed from the TOAC oxidation site. Current attempts in
our laboratories are aimed at designing and synthesizing
chiral TOAC analogues.
crystal system
space group
a[ä]
b [ä]
c [ä]
orthorhombic
P212121
12.120(3)
14.743(3)
19.874(4)
908
908
908
3551(1)
4
1.13
monoclinic
P21
11.702(2)
19.410(3)
12.731(2)
908
108.61(4)8
908
2740(1)
2
a [8]
b [8]
g [8]
V [ä3]
Z
1calcd [MgmÀ3
]
1.15
0.62
1026
m [mmÀ1
F(000)
]
0.61
1308
crystal size [mm]
q range [8]
index ranges
0.30 Â 0.25 Â 0.20
3.73 59.99
À 1 ꢁ h ꢁ 13
0 ꢁ k ꢁ 16
0 ꢁ l ꢁ 22
3282
3244 [R(int) ˆ 0.030]
3244/5/384
0.94
0.40 Â 0.30 Â 0.20
3.66 59.99
À 13 ꢁ h ꢁ 12
0 ꢁ k ꢁ 21
0 ꢁ l ꢁ 14
4409
4203 [R(int) ˆ 0.049]
4198/9/649
1.01
reflections collected
independent reflections
data/restraints/parameters
goodness-of-fit (on F 2)
final R indices [I > 2s(I)]
R indices (all data)
R1 ˆ 0.040, wR2 ˆ 0.099 R1 ˆ 0.038, wR2 ˆ 0.101
R1 ˆ 0.060, wR2 ˆ 0.109 R1 ˆ 0.047, wR2 ˆ 0.106
largest diff. peak/hole [eäÀ3
]
0.166/ À 0.114
0.185/ À 0.158
The aromatic rings of the Fmoc moiety of dipeptide amide (1) were
constrained to the idealized geometry and restraints were applied to the
À
C C bond lengths of the C-terminal tert-butyl moiety. The atoms of the
C-terminal tert-butyl group of pentapeptide amide (15) were refined on two
sets of positions (CT1, CT2, CT3, CT4 and CT1', CT2', CT3', CT4',
respectively) with a population parameter of 0.50 for each set.
Crystallographic data (excluding structure factors) for the structures
reported in this paper have been deposited with the Cambridge Crystallo-
graphic Data Centre as supplementary publication no. CCDC 163805 and
CCDC 163806. Copies of the data can be obtained free of charge on
application to CCDC, 12 Union Road, Cambridge CB2 1EZ, UK CB21EZ,
(fax: (‡44)1223-336-033; e-mail: deposit@ccdc.cam.ac.uk).
Chemical oxidation: The experimental procedure was that described by
Anelli et al.[38] The oxidation was performed by using the NaOCl/KBr
system with 1.5 mol equivalents of the nitroxyl catalyst in a rapidly stirred,
two-phase CH2Cl2/H2O (pH 8.6) mixture at 08C. The efficiency of the
resolution was given by the selectivity factor S ˆ ln[(1 À C)(1 À ee)]/
ln[(1 À C)(1‡ee)], in which ee is the fractional enantiomeric excess and C
is the conversion.[58]
Experimental Section
Materials: The physical properties and analytical data for the new TOAC
and (aMe)Val derivatives and peptides are listed in Table 1. The synthesis
and characterization of Fmoc-[l-(aMe)Val]n-TOAC-[l-(aMe)Val]2-
NHtBu (15: n ˆ 2; 16: n ˆ 3) have already been reported.[11]
FTIR spectroscopy: FTIR absorption spectra were recorded with
a
Perkin Elmer1720X FTIR spectrophotometer, nitrogen flushed, with a
sample-shuttle device, at 2 cmÀ1 nominal resolution, averaging 100 scans.
Solvent (baseline) spectra were recorded under the same conditions. Cells
with CaF2 windows and path lengths of 0.1, 1.0 and 10 mm were used.
2
Spectrograde [2H]chloroform (99.8% H) was obtained from Fluka.
Quantitative analyses of 1-phenylethanol and acetophenone were carried
out with a Varian model3700 gas chromatographic (GC) apparatus based
on a 20-M 15% Carbowax column supported on Chromosorb WAW-
DMCS. 1-Phenylethanol ee was determined by GC analysis with a C. Erba
model HRGC apparatus based on a Chiraldex GTA (30 m  0.32 mm)
column.
X-ray diffraction: Crystals of Fmoc-TOAC-l-(aMe)Val-NHtBu (1) and
Fmoc-[l-(aMe)Val]2-TOAC-[l-(aMe)Val]2-NHtBu (15) were grown by
slow evaporation from ethyl acetate/petroleum ether and methanol,
respectively. Diffraction data were collected on
a Philips PW1100
diffractometer. Crystallographic data are summarized in Table 7.
Both structures were solved by direct methods with the SHELXS97[54]
or SHELXS96[55] programs. Refinements were carried out on F 2
by full-matrix block least-squares, using all data, by application
of the SHELXL97[56] and SHELXL93[57] programs, respectively,
with all non-H atoms anisotropic and allowing their positional parameters
and the anisotropic displacement parameters to refine at alternate
cycles.
Electrochemical oxidation: The instrumentation employed for the electro-
chemical measurements was an EG&G-PARC173/179 potentiostat-digital
coulometer. Electrochemical measurements were performed in an un-
divided glass cell, with thermostatic control at the required temperature.
The cell was equipped with a thermometer and two Pt electrodes. The
working electrode was a Pt plate, having a surface area of 2 cm2, whereas
the counter electrode was a large surface Pt grid. Argon was continuously
bubbled into the aqueous electrolytic mixture during the experiments to
facilitate the removal of hydrogen evolved at the cathode.
H atoms of both compounds were calculated at idealized positions. During
the refinement they were allowed to ride on their carrying atom with Uiso
set equal to 1.2 (or 1.5 for methyl groups) times the Ueq of the parent atom.
Chem. Eur. J. 2002, 8, No. 1
¹ WILEY-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002
0947-6539/02/0801-0091 $ 17.50+.50/0
91

C. Toniolo et al.
FULL PAPER
Typically, the experiments were carried out according to the following
procedure: A mixture of racemic 1-phenylethanol (0.83 mmol) and the
nitroxyl mediator (0.014 mmol) was dissolved in CH2Cl2 (3.5 mL) and
aqueous NaBr (9 mL, 20w%), saturated with NaHCO3, was added. The
two Pt electrodes were immersed in the upper aqueous layer of the
resulting two-phase solution, and the mixture was electrolyzed under a
constant current flow of 20 mA with moderate magnetic stirring. The
electrolysis was interrupted after consumption of 2 FmolÀ1 (usually, after
ca. 2 h) and the mixture was stirred for a further hour to allow complete
consumption of the reactive species. Reaction mixture work-up, quantita-
tive analysis and ee-determination were carried out as described above for
the chemical oxidation.
[18] W. H. Kruizinga, J. Bolster, R. M. Kellogg, J. Kamphuis, W. H. J.
Boesten, E. M. Meijer, H. E. Schoemaker, J. Org. Chem. 1988, 53,
1826 1827.
[19] T. Sonke, B. Kaptein, W. H. J. Boesten, Q. B. Broxterman, H. E.
Schoemaker, J. Kamphuis, F. Formaggio, C. Toniolo, F. P. J. T. Rutjes
in Stereselective Biocatalysis (Ed.: R. N. Patel), Dekker, New York,
1999, pp. 23 58.
[20] L. A. Carpino, E. S. M. E. Mansour, D. Sadat-Aalaee, J. Org. Chem.
1991, 56, 2611 2614.
[21] R. Marchetto, S. Schreier, C. R. Nakaie, J. Am. Chem. Soc. 1993, 115,
11042 11043.
[22] C. R. Nakaie, S. Schreier, A. C. M. Paiva, Biochim. Biophys. Acta
1983, 742, 63 71.
[23] L. Dulog, W. Wang, Liebigs Ann. Chem. 1992, 301 303.
[24] L. A. Carpino, J. Am. Chem. Soc. 1993, 115, 4397 4398.
[25] G. M. Bonora, C. Mapelli, C. Toniolo, R. R. Wilkening, E. S. Stevens,
Int. J. Biol. Macromol. 1984, 6, 179 188.
[26] M. Palumbo, S. Da Rin, G. M. Bonora, C. Toniolo, Makromol. Chem.
1976, 177, 1477 1492.
[27] G. Valle, G. M. Bonora, C. Toniolo, Can. J. Chem. 1984, 62, 2661
2666.
[28] P. Chakrabarti, J. D. Dunitz, Helv. Chim. Acta 1982, 65, 1555 1562.
[29] E. Benedetti in Chemistry and Biochemistry of Amino Acids, Peptides,
and Proteins, Vol. 6 (Ed.: B. Weinstein), Dekker, New York, 1982,
pp. 105 184.
[30] T. Ashida, Y. Tsunogae, I. Tanaka, T. Yamane, Acta Crystallogr. Sect.
B 1987, 43, 212 218.
[31] J. L. Flippen-Anderson, C. George, G. Valle, E. Valente, A. Bianco, F.
Formaggio, M. Crisma, C. Toniolo, Int. J. Pept. Protein Res. 1996, 47,
231 238.
[32] M. Crisma, V. Monaco, F. Formaggio, C. Toniolo, C. George, J. L.
Flippen-Anderson, Lett. Pept. Sci. 1997, 4, 213 218.
[33] IUPAC IUB Commission on Biochemical Nomenclature, Biochem-
istry 1970, 9, 3471 3479.
Conformational energy calculations: The conformational spaces for the
diastereomeric intermediates IV (Scheme 1) between (R) [and (S)]-1-
phenylethanol and Fmoc-TOAC-l-(aMe)Val-NHtBu were mapped by
À
calculating the energy at 58 intervals for the torsion angles around the N O
À
and O C bonds. Minimum-energy conformations were obtained in the low-
energy regions located in the above search, minimizing the energy with
respect to all geometrical parameters, by using the conjugate algorithms.
The structure of Fmoc-TOAC-l-(aMe)Val-NHtBu used in the computa-
tional analysis was that previously obtained from the X-ray diffraction
analysis described in this paper. The geometry of the acetamido group of
the reference peptide Ac-l-Ala-l-Ala-NHtBu was that proposed by
Scheraga and co-workers.[59] The geometrical parameters for the l-Ala
residue were those reported in the InsightII/Biopolymers module.[60]
Energy minimization calculations were performed with the Insight/
DISCOVER package[60] with the Consistent Valence Force Field
63]
(CVFF).[61 Minimum-energy conformations were obtained by minimiz-
ing the energy with respect to all geometrical parameters using the
conjugate algorithms. All conformational energies are expressed as DE ˆ
E À E0, in which E0 is the energy of the most stable conformation. All
computations were performed on a Silicon GraphicsO2 workstation of the
Biocrystallography Research Centre, CNR, at the University of Naples
™FedericoII∫.
[34] C. Ramakrishnan, N. Prasad, Int. J. Protein Res. 1971, 3, 209 231.
[35] R. Taylor, O. Kennard, W. Versichel, Acta Crystallogr. Sect. B 1984, 40,
280 288.
[36] C. H. Gˆrbitz, Acta Crystallogr. Sect. B 1989, 45, 390 395.
[37] D. Cremer, J. A. Pople, J. Am. Chem. Soc. 1975, 97, 1354 1358.
[38] P. L. Anelli, C. Biffi, F. Montanari, S. Quici, J. Org. Chem. 1987, 52,
2559 2562.
[1] C. M. Venkatachalam, Biopolymers 1968, 6, 1425 1436.
[2] C. Toniolo, Crit. Rev. Biochem. 1980, 9, 1 44.
[3] G. D. Rose, L. M. Gierasch, J. A. Smith, Adv. Protein Chem. 1985, 37,
1
109.
[39] J. M. Bobbit, M. C. L. Flores, Heterocycles 1988, 27, 509 533.
[40] S. D. Rychnovski, R. Vaidyanathan, T. Beauchamp, R. Lin, P. J.
Farmer, J. Org. Chem. 1999, 64, 6745 6749.
[41] Z. Ma, Q. Huang, J. M. Bobbitt, J. Org. Chem. 1993, 58, 4837 4843.
[42] S. D. Rychnovski, T. L. Mc Lernon, H. Rajapakse, J. Org. Chem. 1996,
61, 1194 1195.
[43] N. Naik, R. Braslau, Tetrahedron 1998, 54, 667 696.
[44] S. D. Rychnovski, T. Beauchamp, R. Vaidyanathan, T. Kwan, J. Org.
Chem. 1998, 63, 6363 6374.
[45] Y. Kashiwagi, Y. Yanagisawa, F. Kurashima, J. Anzai, T. Osa, J. M.
Bobbitt, Chem. Commun. 1996, 2745 2746.
[4] C. Toniolo, E. Benedetti, Trends Biochem. Sci. 1991, 16, 350 353.
[5] I. L. Karle, P. Balaram, Biochemistry 1990, 29, 6747 6756.
[6] C. Toniolo, E. Benedetti, Macromolecules 1991, 24, 4004 4009.
[7] C. Toniolo, M. Crisma, F. Formaggio, Biopolymers 1998, 47, 153 158.
[8] C. Toniolo, E. Valente, F. Formaggio, M. Crisma, G. Pilloni, C.
Corvaja, A. Toffoletti, G. V. Martinez, M. P. Hanson, G. L. Millhauser,
C. George, J. L. Flippen-Anderson, J. Pept. Sci. 1995, 1, 45 57.
[9] V. Monaco, F. Formaggio, M. Crisma, C. Toniolo, P. Hanson, G.
Millhauser, C. George, J. R. Deschamps, J. L. Flippen-Anderson,
Bioorg. Med. Chem. 1999, 7, 119 131.
[10] C. Toniolo, F. Formaggio, M. Crisma, J. P. Mazaleyrat, M. Wakselman,
C. George, J. R. Deschamps, J. L. Flippen-Anderson, B. Pispisa, M.
Venanzi, A. Palleschi, Chem. Eur. J. 1999, 5, 2254 2264.
[46] Y. Kashiwagi, F. Kurashima, C. Kikuchi, J. Anzai, T. Osa, J. M.
Bobbitt, Tetrahedron Lett. 1999, 40, 6469 6472.
[47] Y. Kashiwagi, K. Uchiyama, F. Kurashima, C. Kikuchi, J. Anzai,
Chem. Pharm. Bull. 1999, 47, 1051 1052.
[48] E. M. Belgsir, H. J. Sch‰fer, Chem. Commun. 1999, 435 436.
[49] M. Kuroboshi, H. Yoshihisa, M. N. Cortona, Y. Kawakami, Z. Gao, H.
Tanaka, Tetrahedron Lett. 2000, 41, 8131 8135.
[11] B. Pispisa, C. Mazzuca, A. Palleschi, L. Stella, M. Venanzi, F.
Formaggio, A. Polese, C. Toniolo, Biopolymers 2000, 55, 425 435.
[12] G. Valle, M. Crisma, C. Toniolo, S. Polinelli, W. H. J. Boesten, H. E.
Schoemaker, E. M. Meijer, J. Kamphuis, Int. J. Pept. Protein Res. 1991,
37, 521 527.
[50] H. Tanaka, Y. Kawakami, K. Goto, M. Kuroboshi, Tetrahedron Lett.
2001, 42, 445 448.
[51] T. Inoguchi, S. Matsumoto, S. Torii, J. Org. Chem. 1991, 56, 2416 2421.
[52] C. A. Hunter, K. R. Lawson, J. Perkins, C. J. Urch, Perkin 2 2001,
651 669.
[13] C. Toniolo, M. Crisma, G. M. Bonora, B. Klajc, F. Lelj, P. Grimaldi, A.
Rosa, S. Polinelli, W. H. J. Boesten, E. M. Meijer, H. E. Schoemaker,
J. Kamphuis, Int. J. Pept. Protein Res. 1991, 38, 242 252.
[14] F. Formaggio, M. Pantano, G. Valle, M. Crisma, G. M. Bonora, S.
Mammi, E. Peggion, C. Toniolo, W. H. J. Boesten, H. E. Schoemaker,
J. Kamphuis, Macromolecules 1993, 26, 1848 1852.
¬
[53] N. Moitessier, B. Maigret, F. Chretien, Y. Chapleur, Eur. J. Org. Chem.
2000, 995 1005.
[15] C. Toniolo, M. Crisma, F. Formaggio, G. Valle, G. Cavicchioni, G.
¬
[54] G. M. Sheldrick, SHELXS97, Program for the Solution of Crystal
Structures, University of Gˆttingen, Gˆttingen, Germany, 1997.
[55] G. M. Sheldrick, SHELXS96, Program for the Solution of Crystal
Structures, University of Gˆttingen, Gˆttingen, Germany, 1996.
[56] G. M. Sheldrick, SHELXL97, Program for the Refinement of Crystal
Structures, University of Gˆttingen, Gˆttingen, Germany, 1997.
Precigoux, A. Aubry, J. Kamphuis, Biopolymers 1993, 33, 1061 1072.
[16] A. Polese, F. Formaggio, M. Crisma, G. Valle, C. Toniolo, G. M.
Bonora, Q. B. Broxterman, J. Kamphuis, Chem. Eur. J. 1996, 2, 1104
1111.
[17] C. Toniolo, A. Polese, F. Formaggio, M. Crisma, J. Kamphuis, J. Am.
Chem. Soc. 1996, 118, 2744 2745.
92
¹ WILEY-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002
0947-6539/02/0801-0092 $ 17.50+.50/0
Chem. Eur. J. 2002, 8, No. 1

Nitroxyl Peptides as Catalysts
84 93
[57] G. M. Sheldrick, SHELXL93, Program for the Refinement of Crystal
Structures, University of Gˆttingen, Gˆttingen, Germany, 1993.
[58] H. B. Kagan, J. C. Fiaud, Top. Stereochem. 1988, 18, 249 330.
[61] S. Lifson, A. T. Hagler, P. J. Dauber, J. Am. Chem. Soc. 1979, 101,
5111 5121.
[62] A. T. Hagler, S. Lifson, P. J. Dauber, J. Am. Chem. Soc. 1979, 101,
¬
[59] G. Nemethy, M. S. Pottle, H. A. Scheraga, J. Phys. Chem. 1983, 87,
5122 5130.
1833 1887.
[63] A. T. Hagler, P. J. Dauber, S. Lifson, J. Am. Chem. Soc. 1979, 101,
[60] InsightII and Discover User×s Guides, Molecular Simulations, San
Diego, CA, 2000.
5131 5140.
Received: July 13, 2001 [F3413]
Chem. Eur. J. 2002, 8, No. 1
¹ WILEY-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002
0947-6539/02/0801-0093 $ 17.50+.50/0
93