Protein secondary structure mimetics: Crystal conformations of α/γ4-hybrid peptide12-helices with proteinogenic side chains and their analogy with α- And β-peptide helices

Article abstract of DOI:10.1039/c2ob26805a

Numerous strategies have been developed to mimic the α-helical secondary structure using hybrid peptides containing non-natural amino acids. In contrast to the β- and α/β-hybrid peptides, very little is known about the folding patterns of hybrid peptides

Full text of DOI:10.1039/c2ob26805a

Organic &
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Protein secondary structure mimetics: crystal
conformations of α/γ4-hybrid peptide12-helices with
proteinogenic side chains and their analogy with
α- and β-peptide helices†
Cite this: Org. Biomol. Chem., 2013, 11,
509
Sandip V. Jadhav, Anupam Bandyopadhyay and Hosahudya N. Gopi*
Numerous strategies have been developed to mimic the α-helical secondary structure using hybrid pep-
tides containing non-natural amino acids. In contrast to the β- and α/β-hybrid peptides, very little is
known about the folding patterns of hybrid peptides containing γ4-amino acids. Here we report the solid
phase synthesis and crystallographic insight into the secondary structures formed by 1 : 1 alternating
α/γ4-hybrid peptides. The crystal conformations suggest that heptapeptides P1, P2 and P3 adopted the
12-helix conformation with backward consecutive 1←4 H-bonds [CvO(i)⋯H–N (i + 3)]. In comparison
with α-, β- and γ-peptides, the distinct projection of side-chains was observed along the helical cylinder.
In contrast to the peptide containing stereochemically constrained α-amino acid Aib (P1), the peptide
with complete proteinogenic side-chains (P3) displayed organized side chain–side chain interactions
between the antiparallel helices in crystal packing. The analogy of the α/γ4-hybrid peptides with
310-helix, α-helix and β-peptide 12-helix suggests that the internal H-bonding pattern and macrodipole
were analogous to the α- and β-peptide helices. In addition, helical parameters were found to be very
similar to that of β-peptide 12-helices.
Received 14th September 2012,
Accepted 19th November 2012
DOI: 10.1039/c2ob26805a
www.rsc.org/obc
structures.5 The advantage of hybrid peptides with a hetero-
geneous backbone (peptides with α and other β- and γ-amino
Introduction
Helices constitute the major secondary structural components acids) is that a variety of hydrogen bonded helical structures
of proteins and often play a crucial role in mediating protein– can be generated by varying the amino acid sequence patterns.
protein and protein–nucleic acid (DNA and RNA) interactions.1 Extensive investigations revealed that β-, γ-, mixed α/β- and
Disrupting these interactions with isolated helical structures is α/γ-hybrid peptides adopt various ordered helical confor-
of paramount importance not only to understand the biologi- mations including 14-, 12-, 10/12-, 9- and 8-helices. In contrast
cal consequences of the interactions but also from the perspec- to the 14-helical conformations of cyclic β-amino acids with a
tive of drug design. Several approaches have been developed to six membered ring constraint (trans-2-aminocyclohexane car-
6
mimic the short and stable α-peptide helices including boxylic acid) and acyclic β-amino acids (β3- and β2-amino
covalent cross-linkage of amino acid side-chains,2 utilization acids),5a the homooligomers of cyclic β-amino acids with four
of C–C covalent bonds as an intramolecular H-bond surrogate 3 (trans-2-aminocyclobutane carboxylic acid),7 five (trans-2-amino-
or non-peptidic organic templates.4 The recent exponential cyclopentane carboxylic acid and its derivatives)8 membered
growth of β- and γ-peptide foldamers provides an alternative ring constraint and a bicyclo[2.2.1] heptene skeleton9 dis-
and straightforward approach to mimic protein secondary played the 12-helical conformations. The cyclic ring con-
straints of these amino acids preorganize the β-peptide
backbone to adopt helical conformations. In addition,
Department of Chemistry, Indian Institute of Science Education and Research,
Gellman et al.10 and others11 have demonstrated the inhi-
Dr. Homi Bhabha Road, Pune-411 008, India. E-mail: hn.gopi@iiserpune.ac.in
bitions of protein–protein interactions using 12-helical
scaffolds. Further, Lee et al. showed the exceptional self-assem-
bly behaviour of the 12-helical peptides.12 Recent theoretical
results suggest that the 12-helical conformation is not unique
to the homooligomers of cyclic β-amino acids, the heterooligo-
mers composed of 1 : 1 α- and unsubstituted γ-amino acids
†Electronic supplementary information (ESI) available: Details of chemical
synthesis, characterization and crystallographic details of γ4-Phe and peptides
P1, P2 and P3, average helical parameter table, hydrogen bond parameters and
torsional angle tables, HPLC traces, CD analysis and complete spectroscopic
data for all compounds. CCDC . CCDC reference numbers 901427–901430. For
ESI and crystallographic data in CIF or other electronic format see DOI: 10.1039/
c2ob26805a
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have shown to adopt 12-helix.13 In their experimental work,
Balaram and colleagues14 and Gellman et al.15 have shown
12-helical conformations in α/γ-hybrid peptides containing
acyclic γ-amino acids and cyclic γ-amino acids, respectively.
Recently, we have shown the 12-helical organizations in the
short α/γ4-hybrid peptides obtained through the direct trans-
formation from α/vinylogous hybrid peptides using catalytic
hydrogenation.16 In all these cases stereochemically con-
strained amino acids have been used to induce the helical con-
formation in the peptides. As proteinogenic amino acid side
chains play a pivotal role in biomolecular interactions, we
sought to investigate structural properties of α/γ4-hybrid pep-
tides with complete proteinogenic side chains. Herein, we are
reporting the solid phase synthesis, single crystal confor-
mations of α/γ4-hybrid heptapeptides Ac-Aib-γ4Phe-Aib-γ4Phe-
Aib-γ4Phe-Aib-CONH2 (P1), Ac-Ala-γ4Phe-Ala-γ4Phe-Ala-γ4Phe-
Aib-CONH2 (P2), Ac-Ala-γ4Phe-Ala-γ4Phe-Ala-γ4Phe-Ala-CONH2
(P3) and their analogy with 310-helix, α-helix and β-peptide
12-helices.
Fig. 1 X-ray structures of free γ4-Phe. Conformation and torsional values are
given below.
Results and discussion
Design and synthesis of α/γ4-hybrid peptides
To understand whether α/γ4-hybrid peptides can fold into
helical conformations without any stereochemical constraint,
we designed three heptapeptides P1, P2 and P3. We have uti-
lised γ4-Phe in combination with α amino acids to synthesize
α/γ4-hybrid peptides. The solid phase compatible Fmoc-γ4-Phe
was synthesized through the catalytic hydrogenation of the
benzyl ester of N-Cbz-protected α, β-unsaturated γ-phenyl-
alanine followed by the Fmoc-protection as shown in
Scheme 1. In addition, single crystals of free amino acid γ4-
Phe obtained as an intermediate in the process of the reaction
yield the structure shown in Fig. 1. The advantage of this
method is that the γ4-amino acids can be directly incorporated
into the peptide sequence using solid phase synthesis. The
stereochemically constrained helix favouring α,α-dialkyl amino
acid (Aib)17 was used as an α-amino acid in 1 : 1 alternating
α/γ4-hybrid peptide P1. In the case of P2, except the C-terminal
Aib, all other Aibs are replaced with L-Ala. In P3 all Aibs are
Scheme 2 α/γ4-Hybrid peptides P1, P2 and P3 synthesized using solid phase
peptide synthesis.
replaced with L-Ala. The sequences of these peptides are
shown in Scheme 2. All peptides were synthesized on a Knorr
Amide MBHA resin using the standard Fmoc-chemistry proto-
col. The coupling reactions were mediated by HBTU/HOBt
coupling conditions.
Crystal structures are very important in understanding the
helical parameters such as residue-per-turn, rise-per-turn,
helical radius and the projection of amino acid side-chains in
the newly designed peptides. Keeping this in mind, we
attempted to grow the X-ray quality crystals from all three
peptides in various solvent combinations.
Crystal conformations of α/γ4-hybrid peptides
Single crystals of peptides P1 and P2 were obtained from the
slow evaporation of aqueous methanol–trifluoroethanol solu-
tion. The suitable X-ray quality single crystals of P3 were
obtained after repeated attempts of slow evaporation from
aqueous methanol solution. The X-ray structures of all three
peptides are shown in Fig. 2. Instructively, all three peptides
adopted right-handed helical conformations with consecutive
Scheme 1 (a) Synthesis of Fmoc-γ4-Phe and (b) local conformational variables
of γ4-amino acids.
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Fig. 3 (a) A two dimensional Ramachandran type plot depicting ϕ and ψ
values in the α/γ4-hybrid peptides. The torsional variables θ1 and θ2 are kept
constant as they always take the values close to 60° in the hybrid helices. (b) The
helical wheel diagram depicting side chain projection in α/γ4-hybrid peptides.
α-Amino acids are shown in red while γ4-amino acids are shown in green.
conformations about the Cβ–Cγ and Cα–Cβ bonds. The average
ϕ and ψ values of γ4-residues were found to be 125 7 and
−118 10°, respectively. The H-bonding pattern between the
1←4 residues and directionality of the H-bond in all α/γ4
hybrid peptides (P1–P3) indicating the backbone expanded
version of a 310-helix.17b,21 In contrast to the average ϕ and ψ
values of the 310-helix (−49°, −26°), L-Ala in the peptides P2
and P3 displayed the average ϕ and ψ values −68 3° and
−33.03 3°, respectively. The torsional angles of all hybrid
peptides are tabulated and given in the ESI.†. A plot of ϕ and ψ
angles of all residues in peptides P1–P3 is shown in Fig. 3a.
Keeping θ1 and θ2 as constants, two distinct regions in the left
quadrant of the Ramachandran map18 can be recognised for
γ- and α-residues.
Fig. 2 (a) X-ray structures of P1, P2 and P3. Top view is shown in the lower
plane. (b) The lateral interaction of anti-parallel 12-helices of P3 in the crystal
packing and the top view of 12-helix P3 are also shown.
In comparison between the α-residues in α/γ4-hybrid pep-
tides and 310-helix, a clear distinction can be observed particu-
larly in the ϕ values. The L-Ala displayed an average of 10°
higher value over the sterically constrained dialkyl amino acids
12-membered H-bonds [CvO(i)⋯H–N(i + 3), 12-atom ring in α/γ4-hybrid peptides and an average of 19° over the α-resi-
H-bonds]. Similar to the native α-amino acid helices, dues in the 310-helix. The crystal structure results suggest that
α/γ4-hybrid helices displayed the backward H-bonding direc- a stable 12-helix can be constructed using either the stereoche-
tionality and subsequent macrodipole. The 12-helical confor- mically constrained Aib or simple unconstrained α-residues in
mation in all the α/γ4-hybrid peptides is stabilized by the combination with alternating γ4-residues. Further, the top
six consecutive 1←4 [CvO(i)⋯H–N(i + 3)] intramolecular view of the 12-helices indicates the projections of the side
H-bonds. Both a C-terminal amide and an N-terminal Ac-group chains at four corners of the helical cylinder (Fig. 2a, bottom
are involved in the intramolecular H-bonds. Additionally, the plane). In comparison to the α-, β- and γ-peptides, the distinct
packing mode of individual peptides revealed that each helical orientation of the amino acid side-chains was observed in α/γ4-
peptide is interconnected with the other helical peptides in a hybrid 12-helices. Based on this information we predicted the
head-to-tail fashion through four intermolecular H-bonds. The helical wheel diagram for α/γ4-hybrid peptide 12-helices
inter- and intramolecular H-bond parameters of all three pep- (Fig. 3b). In hindsight, the analysis of the packing modes of all
tides are tabulated in the ESI.† Inspection of the crystal struc- three peptide helices reveals the importance of proteinogenic
ture of P1 reveals that Aib residues adopted right handed amino acid side-chains. In contrast to P1 and P2, P3 with
helical conformations by having average ϕ and ψ values −58 3° α-alanine residues showed the lateral interactions between the
and −40
5°, respectively. The dihedral angles of γ4-Phe two anti-parallel helices. The packing of antiparallel helices is
residues were measured by introducing two additional vari- stabilized by the side-chain hydrophobic as well as CH⋯π
ables θ1 (N–Cγ–Cβ–Cα) and θ2 (Cγ–Cβ–Cα–C) as shown in interactions. The anti-parallel packing of P3 is shown in
Scheme 1b. In contrast to the amino acid structure, the stereo- Fig. 2b. These packing interactions may provide the guidance
chemical analysis of γ4-Phe residues in all the peptides reveals for structure based design from α/γ4-hybrids to target protein
that they adopted gauche+, gauche+ (g+, g+, θ1 ≈ θ2 ≈ 60°) local structures. In addition two water molecules located at C- and
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the β-peptide8 12-helix is shown in Fig. 4a. For comparison,
the average structural parameters of β-peptide 12-helices and
the 310-helix21 are tabulated in Table 1. In addition, α/γ4-
hybrid peptide 12-helices displayed a very similar CD signature
as that of β-peptide 12-helices with a CD maximum at 205 nm
and a weak minimum at 218 nm (See ESI†).20 The negative para-
meters of 12-helices generated from the cyclobutane ring con-
straints confirm the left-handed helical conformation of the
β-peptide.7 Further, we superimposed the α/γ4-hybrid peptide
P3 over the α-helix and the 310-helix to understand the
backbone correlation and the side-chain orientation as the
H-bonding directionality of hybrid peptide 12-helices was very
similar to that of α-peptide helices. The superposition of the
backbone conformations of P3 over the α-helix (from the
protein Human DNA Polymerase Beta, PDB code – 1ZQA,
sequence – 94–102)22 and the 310-helix23 is shown in Fig. 4b
and 4c respectively. Similar to the α, γ4-hybrid hexapeptide,16b
the backbone conformation of the hybrid heptapeptide P3 is
well correlated with the nine residues of the α-helix. Instruc-
tively, a good backbone correlation of heptapeptide P3 was
observed with the heptapeptide 310-helix. The structures of the
β-peptide 12-helix, α-helix and 310-helix were generated using
the co-ordinates reported in the literature.24 The top view of
the superimposed P3 with the α-helix and the 310-helix sig-
nifies the projection of the amino acid side-chains (Fig. 4,
down plane). Similar results were also observed for the hybrid
peptides P1 and P2 (data not shown). The backbone corre-
lation and the side-chain projections of α/γ4-hybrid peptide
helices with respect to the α-helix suggest that these hybrid
peptides can be exploited as mimics of α-peptide helices.
Further, with the availability of broad side-chain diversity in
both α- and γ4-amino acids, these α/γ4-hybrid peptides stand
Table 1 Average helical parameters of backward 1←4 H-bonded helices
Res/
turn
n
Rise/
turn
p (Å)
Rise/
res
d (Å)
Radius
r (Å)
Peptide backbone
α/γ4-Peptide (12-helix)
β-Peptide(trans ACPC)12-
helixa
2.7
2.7
5.3
5.4
2.0
2.0
2.1
2.1
β-Peptide(trans ACBC)12-
−2.7d
−5.4d
2.0
2.2
helixb
310-Helixc
3.2
5.8
1.8
2.0
a Ref. 8. b Ref. 7. c Ref. 21. d Negative sign indicates the left handed
helical turn.
Fig. 4 Superposition of α/γ4-hybrid peptide P3 (light blue) on (a) β-peptide
12-helix, (b) α-helix and (c) 310-helix. The top view (lower plane) depicting the unique from the other α/γ-hybrid peptides. The structural
projection of amino acid side-chains.
analysis of α/γ-hybrid peptides containing backbone homolo-
gated γ4-amino acids with proteinogenic side chains presented
here may be useful for the design of functional foldamers
similar to the β- and α/β-hybrid peptides.
N-terminals of the antiparallel helices play a crucial role in
interconnecting the two helices.
Structural analogy with α-peptide helices, β-peptide 12-helices
and helical parameter analysis
Conclusion
The intriguing results from the hybrid peptides P1, P2 and P3 In conclusion, we have presented the facile solid phase syn-
encourage us to determine the helical parameters of α/γ4- thesis and single crystal conformations of α/γ4-hybrid pep-
hybrid peptide 12-helices. The helical parameters were calcu- tides. The stereochemical analysis suggests that all three α/γ4-
lated from the set of four consecutive α-carbons using reported hybrid heptapeptides adopted 12-helical conformations in
methods.19 The analysis reveals all characteristic features such single crystals. Comparison between the P1 and P3 reveals that
as residue-per-turn, rise-per-turn and the radius of α/γ4-hybrid the 12-helical conformation can be induced in a hybrid
peptide 12-helix. The average helical parameters obtained peptide sequence solely through intramolecular H-bonding
from the three crystal structures are tabulated in Table 1. without using any stereochemical constraints. In addition,
Recently, Gellman et al. reported the helical parameters for the analogy with the β-peptide 12-helix suggests that α/γ4-hybrid
β-peptide 12-helix8 containing stereochemically constrained peptides can be used as surrogates of β-peptide 12-helices.
cyclic β-amino acids. Interestingly, the helical parameters cal- Analysis of the helical parameters and the backbone corre-
culated for the α/γ4-hybrid peptides are in good agreement lation with the 310-helix suggests that the α/γ4-hybrid peptide
with the β-peptide12-helices [residue-per-turn (2.7), rise- is comparable to an α-peptide helix except the projection of
per-turn (5.4 Å) and radius (2.1 Å)] generated from the cyclo- the amino acid side chains. The distinct location of the amino
pentane backbone constraints. The overlay of the peptide P3 on acid residues in the helical wheel diagram presented here may
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be useful in the design of hybrid peptides with specific reflections. Space group P21; a = 7.757(5), b = 6.483(4),
patterns. We believe that the facile protocol for the synthesis c = 9.907(7) Å; α = γ = 90.00°, β = 92.808(12) (3)°. V = 497.6(6)
of γ4-amino acids, crystal conformations of the hybrid peptides Å3, monoclinic P; Z = 2 for chemical formula C11H15N1O2;
and their analogy with α- and β-peptide helices presented here ρcalcd = 1.290 Mg m−3; μ = 0.088 mm−1; F (000) = 208. The
may offer the guidelines to design α/γ4-hybrid peptides with structure was obtained by direct methods using SHELXS-97.
specific functions.
All non-hydrogen atoms were refined anisotropically. The
hydrogen atoms were fixed geometrically in the idealized posi-
tion and refined in the final cycle of refinement as riding over
the atoms to which they are bonded. The final R value was
0.0704 (wR2 = 0.1626) for 2196 observed reflections (F0 ≥ 4σ
(|F0|)) and 128 variables; S = 0.947. The largest difference peak
and hole were 0.424 and −0.314 e Å−3, respectively.
Experimental
Chemical synthesis of Fmoc-γ4-Phe-OH
The suspension of activated Pd/C (20% by weight) and benzyl
esters of N-Cbz-protected vinylogous phenylalanine (1.66 g,
4 mmol), which was synthesized using the reported method,25
in 10% acetic acid in THF (20 mL) was stirred overnight at
room temperature in the presence of hydrogen. After com-
pletion of the reaction, Pd/C was filtered through the bed of
celite and the filtrate was evaporated to dryness under vacuum
to get gummy free γ4-phenylalanine (γ4-Phe). The pure γ4-Phe
was isolated as a white powder after trituration with cold
diethyl ether in excellent yield (0.687 g, 90%).
Crystal structure analysis of peptide P1. Crystals of α/γ4-
hybrid peptide P1 were grown by slow evaporation from a sol-
ution of aqueous methanol–TFE. A single crystal, rectangular
in shape (0.40 × 0.20 × 0.12 mm3) was mounted on a loop. The
X-ray data were collected at 100(2) K temperature on a Bruker
AXS SMART APEX CCD diffractometer using Mo-Kα radiation
(λ = 0.71073 Å), ω-scans (2θ = 56.50°), for a total number of
26 870 independent reflections. Space group P21; a = 12.866-(15),
b = 17.403(19), c = 13.962(16) Å; α = γ = 90.00°, β = 107.22 (3)°.
V = 2986(6) Å3, monoclinic P; Z = 2 for chemical formula
C51H72N8O8; ρcalcd = 1.029 Mg m−3; μ = 0.07 mm−1; F (000) =
996. The structure was obtained by direct methods using
SHELXS-97. All non-hydrogen atoms were refined anisotropi-
cally. The hydrogen atoms were fixed geometrically in the
idealized position and refined in the final cycle of refinement
as riding over the atoms to which they are bonded. The final
R value was 0.0813 (wR2 = 0.1855) for 13 302 observed reflections
(F0 ≥ 4σ (|F0|)) and 596 variables; S = 0.733. The largest differ-
ence peak and hole were 0.213 and −0.239 e Å−3, respectively.
Crystal structure analysis of peptide P2. Crystals of α/γ4-
hybrid peptide P2 were grown by slow evaporation from a sol-
ution of aqueous methanol–TFE. A single crystal, rectangular
in shape (0.40 × 0.30 × 0.20 mm3), was mounted on a loop.
The X-ray data were collected at 100(2) K temperature on a
Bruker AXS SMART APEX CCD diffractometer using Mo-Kα
radiation (λ = 0.71073 Å), ω-scans (2θ = 56.56°), for a total
number of 24 197 independent reflections. Space group P1;
a = 9.354(10), b = 12.179(13), c = 13.950(15) Å; α = 68.89 (3)°,
β = 85.74 (3)°, γ = 71.36 (3)°, V = 1403(3) Å3, triclinic P; Z = 1 for
Further, to the solution of free γ4-Phe (0.579 g, 3 mmol) in
20% Na2CO3 (15 mL) was added Fmoc-OSu (1.11 g, 3.3 mmol,
dissolved in 10 mL of THF) and the reaction mixture was
stirred overnight. After completion of the reaction, the reaction
mixture was acidified with 10% HCl and the precipitated Fmoc
protected γ4-Phe was extracted with EtOAc (3 × 25 mL). The
combined organic layer was washed with 10% HCl (3 × 15 mL),
brine solution (2 × 10 mL), dried over Na2SO4 and concen-
trated under reduced pressure to give gummy Fmoc-γ4-Phe.
The gummy Fmoc-γ4-Phe was precipitated using ethyl acetate/
pet. ether (60–80 °C) to give 1.06 g (85%) as a white powder
and used directly in the solid phase peptide synthesis.
Peptide synthesis
Peptides P1, P2 and P3 were synthesized by manual solid
phase peptide synthesis on a Knorr Amide MBHA resin by
Fmoc-chemistry (0.25 mmol scale). Coupling reactions were
performed using the HBTU/HOBt/NMP activation protocol.
Fmoc deprotections were facilitated by using 20% piperidine
in DMF. N-terminal of peptides was capped with an acetyl
group. Peptides were cleaved from the resin by using 95% tri-
fluoroacetic acid, 2.5% water and 2.5% triisopropyl silane clea-
vage mixture. Reverse-phase-HPLC purification (detector:
254 nm and 220 nm) on a C18 column (70% methanol to 90%
methanol in 40 min) at a flow rate of 1.5 mL min−1 was carried
out to get pure peptides.
chemical formula C48H66N8O8, (O); ρcalcd = 1.064 Mg m−3
;
μ = 0.074 mm−1; F (000) = 482. The structure was obtained by
direct methods using SHELXS-97. All non-hydrogen atoms
were refined anisotropically. The hydrogen atoms were fixed
geometrically in the idealized position and refined in the final
cycle of refinement as riding over the atoms to which they are
bonded. The final R value was 0.0833 (wR2 = 0.2342) for 10 395
observed reflections (F0 ≥ 4σ (|F0|)) and 569 variables;
Crystal structure analysis
Crystal structure analysis of γ4-Phe. Crystals of γ4-Phe were S = 0.771. The largest difference peak and hole were 0.056 and
grown by slow evaporation from a solution of aqueous metha- −0.257 e Å−3, respectively.
nol. A single crystal, rectangular in shape (0.50 × 0.40 ×
Crystal structure analysis of peptide P3. Crystals of α/γ4-
0.20 mm3) was mounted on a loop. The X-ray data were col- hybrid peptide P3 were grown by slow evaporation from a sol-
lected at 100(2) K temperature on a Bruker AXS SMART APEX ution of aqueous methanol. A single crystal, rectangular in
CCD diffractometer using Mo-Kα radiation (λ = 0.71073 Å), shape (0.40 × 0.29 × 0.16 mm3), was mounted on a loop. The
ω-scans (2θ = 54.88°), for a total number of 8120 independent X-ray data were collected at 100(2) K temperature on a Bruker
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AXS SMART APEX CCD diffractometer using Mo-Kα radiation
(λ = 0.71073 Å), ω-scans (2θ = 56.56°), for a total number of
16 669 independent reflections. Space group P21; a = 11.218(17),
b = 14.58(2), c = 15.75(2) Å; α = γ = 90°, β = 100.06 (3)°, V = 2536(7)
Å3, monoclinic P; Z = 2 for chemical formula C47H64N8O8,
2(O); ρcalcd = 1.140 Mg m−3; μ = 0.079 mm−1; F (000) = 924. The
structure was obtained by direct methods using SHELXS-97.
All non-hydrogen atoms were refined anisotropically. The
hydrogen atoms were fixed geometrically in the idealized posi-
tion and refined in the final cycle of refinement as riding over
the atoms to which they are bonded. The final R value was
0.0824 (wR2 = 0.1713) for 10 827 observed reflections (F0 ≥ 4σ
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Acknowledgements
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