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The crystal structure of L-argininew
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Published on 12 January 2012 on http://pubs.rsc.org | doi:10.1039/C2CC17203H
Emilie Courvoisier,ab P. Andrew Williams,a Gin Keat Lim,a Colan E. Hughesa and
Kenneth D. M. Harris*a
Received 19th November 2011, Accepted 9th January 2012
DOI: 10.1039/c2cc17203h
We report the crystal structure of L-arginine, one of the last
remaining natural amino acids for which the crystal structure
has never been determined; structure determination was carried
out directly from powder X-ray diffraction (XRD) data, exploiting
the direct-space genetic algorithm technique for structure
solution followed by Rietveld refinement.
Of the 20 genetically encoded amino acids found in proteins,
arginine is one of only two cases for which the crystal structure
of a pure crystalline form (either as the single enantiomer or the
racemate) has not yet been reported.1 To date, the only reported
crystal structures containing neutral arginine molecules2 are
3
4
L-arginine dihydrate, DL-arginine monohydrate and DL-arginine
5
dihydrate. The absence of a reported crystal structure for
pure arginine is undoubtedly due to difficulties in obtaining
crystals of sufficient size and quality for single-crystal X-ray
diffraction (XRD) studies. Indeed, our own attempts to crystallize
L-arginine from several different solvents and under a variety of
experimental conditions failed to produce any crystals suitable for
single-crystal XRD.
Although single-crystal XRD is the most powerful experimental
technique for determining crystal structures, the requirement for a
suitable single-crystal specimen imposes a limitation on the applicability of this technique. When a suitable single crystal of the
material of interest cannot be prepared, as in the case encountered
here for L-arginine, structure determination must be tackled instead
from powder XRD data. However, it is important to emphasize
that the task of carrying out structure determination directly from
powder XRD data is considerably more challenging than from
single-crystal XRD data, particularly in the case of organic
materials. Nevertheless, the opportunities for determining the
crystal structures of organic materials from powder XRD data
have advanced considerably in recent years,6 particularly through
the development of the direct-space strategy for structure solution.
In this paper, we exploit these opportunities to determine the
crystal structure of L-arginine directly from powder XRD data.
The powder XRD pattern of L-arginine7 was indexed
using the ITO code8 in the program CRYSFIRE,9 giving the
following unit cell with monoclinic metric symmetry: a = 9.76 Å,
b = 16.02 Å, c = 5.58 Å, b = 98.11 (V = 863.2 Å3). Given the
volume of the unit cell and consideration of density, the
number of molecules in the unit cell was assigned as Z = 4.
From systematic absences, the space group was assigned as
P21 (corresponding to Z 0 = 2) or P21/m (corresponding to
Z 0 = 1). However, as the sample comprises a single enantiomer
of arginine, the achiral space group P21/m is ruled out.
Furthermore, the solid-state 13C NMR spectrum10 provides clear
evidence that the structure contains two crystallographically
independent molecules of L-arginine (Z0 = 2). Hence, the space
group was assigned as P21. Profile fitting using the Le Bail
method11 in the GSAS program12 gave a good quality of fit
(Rwp = 1.71%, Rp = 1.27%; Fig. 1). The refined unit cell and
profile parameters obtained from the Le Bail fit were used in
the subsequent structure-solution calculation.
Structure solution was carried out using the direct-space
genetic algorithm (GA) technique13 incorporated in the program
EAGER.14 In the GA structure-solution calculation, each trial
structure was defined by a total of 25 variables. For one
molecule, the position along the b-axis can be fixed arbitrarily
for space group P21, and thus only two positional variables are
required, while three positional variables are required for the
other molecule; in addition, for each of the two molecules, three
orientational variables and seven torsion-angle variables (Fig. 2)
are required. Hydrogen atoms were included in the structural
a
School of Chemistry, Cardiff University, Park Place,
Cardiff CF10 3AT, Wales, UK. E-mail: HarrisKDM@cardiff.ac.uk
b
Ecole Nationale Supe´rieure de Chimie de Clermont-Ferrand,
Ensemble Scientifique des Ce´zeaux, 24 Avenue des Landais - BP
187, 63174, Aubie`re Cedex, France
w Electronic supplementary information (ESI) available: details of
solid-state 13C NMR spectroscopy. CCDC 855058. For ESI and
crystallographic data in CIF or other electronic format see DOI:
10.1039/c2cc17203h
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Fig. 1 Le Bail fit of the powder XRD pattern of L-arginine (red +
marks, experimental data; green line, calculated data; purple line, difference
plot; black tick marks, predicted peak positions). The high background
arises, at least in part, from scattering by starch, which was mixed with the
7
L-arginine powder to reduce the effects of preferred orientation.
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Fig. 2 Molecular structure of L-arginine showing the seven torsionangle variables in the direct-space structure solution calculation.
model and the specific tautomeric form shown in Fig. 2 was
used, based on the fact15 that the value of pKa is higher for the
guanidinium group (12.48) than the ammonium group (8.99).
We note that this tautomer exists in all other reported crystal
structures containing neutral arginine molecules.2–5
A total of 16 independent GA structure-solution calculations
were carried out. Each calculation involved the evolution of
100 generations for a population of 100 structures, with 40 mating
operations and 30 mutation operations carried out per generation.
Within the relatively small number (100) of generations considered
in these calculations, five calculations converged on essentially the
same structure solution, corresponding to the lowest value of Rwp.
The structure solution (i.e., the structure with lowest Rwp
obtained in the GA calculations) was used as the initial
structural model for Rietveld refinement,16 which was carried
out using the GSAS program.12 Before refinement, inspection
of the structure solution confirmed that the selected tautomeric
form gave a structurally sensible hydrogen-bonding network.
In the Rietveld refinement, standard restraints were applied to
bond lengths and bond angles, planar restraints were applied
to the carboxylate and guanidinium groups, and a global
isotropic displacement parameter was refined. In the latter stages,
a small correction was introduced for preferred orientation. The
final Rietveld refinement (2y range, 4–701; 3867 profile points;
126 refined variables) gave a good fit to the powder XRD data
(Rwp = 1.85%, Rp = 1.33%; Fig. 3), with the following refined
parameters: a = 9.7565(4) Å, b = 16.0230(5) Å, c = 5.5805(3) Å,
b = 98.058(4)1, V = 863.77(10) Å3.
In the crystal structure of L-arginine, the two independent
molecules (denoted types A and B) have very similar conformations (Fig. 4), with an extended side chain lying approximately in
the same plane as the guanidinium group and with the carboxylate
group approximately perpendicular to this plane (interestingly,
this molecular conformation is very similar to that in L-arginine
dihydrate3 but significantly different from those in DL-arginine
monohydrate4 and DL-arginine dihydrate5). The solid-state
13
C NMR spectrum10 is consistent with the two independent
molecules having similar conformations in the crystal structure,
as only two of the six 13C sites in L-arginine exhibit two resolved
Fig. 3 Final Rietveld refinement for L-arginine.
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Chem. Commun., 2012, 48, 2761–2763
Fig. 4 Overlay of the two independent molecules [magenta (type A)
and cyan (type B)] in the crystal structure of L-arginine, with the three
central CH2 groups (grey and white) superimposed directly, demonstrating the very similar molecular conformations.
isotropic peaks as a result of local structural differences for the
two independent molecules.
Along the b-axis, the molecules are arranged in two distinct
‘‘sinusoidal’’ chains (Fig. 5); one chain involves molecules of
type A only and the other chain involves molecules of type B
only. In each chain, the molecules are arranged in a headto-tail manner, and adjacent molecules are linked by two
N–H O hydrogen bonds between the guanidinium (tail)
group of one molecule and the carboxylate (head) group of
the adjacent molecule. These two hydrogen bonds create a
cyclic hydrogen-bonded array with graph set descriptor17
R22(8). Adjacent molecules in a given chain are related by the
21 screw axis. Defining the ‘‘direction’’ of these chains as the
direction of the N–H vector in the N–H O hydrogen bonds,
the chains of molecules of types A and B run in opposite
directions along the b-axis.
Adjacent chains are linked by N–H O hydrogen bonds
between the guanidinium moiety of a molecule in one chain
and the carboxylate groups of molecules in the two neighbouring chains, giving rise to a ribbon motif that extends
along the a-axis (Fig. 6). In addition, as part of this ribbon
motif, the N–H group of the guanidinium moiety of each
molecule is the donor in an N–H N hydrogen bond to a
molecule in a neighbouring chain, with the nitrogen atom of
the amino acid NH2 group as the acceptor. The graph sets of
the three different hydrogen-bonded cycles in these ribbons
are: R22(8) [as discussed above, this same cyclic array links
adjacent molecules in the chains that run along the b-axis],
R24(8) and R22(9). Within a given ribbon, all the hydrogen
bonds lie approximately in the same plane, which is parallel
Fig. 5 Crystal structure of L-arginine viewed along the a-axis, showing only molecules of type A and demonstrating the ‘‘sinusoidal’’
chains that run parallel to the b-axis. Green dashed lines indicate
hydrogen bonds.
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Fig. 6 Crystal structure of L-arginine, viewed perpendicular to the
(041) plane, showing the hydrogen-bonded ribbon motif along the
a-axis (horizontal). Green dashed lines indicate hydrogen bonds.
to either (041) or (041% ). We note that a rather similar hydrogenbonded ribbon motif is present in the crystal structure of
3
L-arginine dihydrate; however, in the dihydrate structure,
the O–H bond of a water molecule is also incorporated within
the hydrogen-bonded array (see ESIw), thus converting the
cyclic array with graph set R24(8) to one with graph set R35(10).
Thus, the overall crystal packing in L-arginine may be
described in terms of severely puckered sheets with an average
plane parallel to the ab-plane; the severe puckering arises from
the sinusoidal topology of the chains that run parallel to the
b-axis. All hydrogen bonding in the structure occurs within
these sheets (i.e., within the chains that run along the b-axis and
the ribbons that run along the a-axis), and stacking of the sheets
along the c-axis involves only van der Waals interactions.
In conclusion, we emphasize that the crystal structure of
L-arginine reported here was determined directly from powder
XRD data, as single crystals of suitable size and quality
for single-crystal XRD studies could not be prepared. Thus,
inter alia, this work serves to demonstrate the opportunities
that now exist for determining the crystal structures of organic
materials using modern techniques for the analysis of powder
XRD data. In more general terms, we also emphasize that
knowledge of the conformational properties and interactions
of individual amino acids in the crystalline state, of the type
reported here, can yield important insights relating to the
structural properties of peptides, and potentially also of polypeptide sequences in proteins.
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12
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14
Notes and references
1 Established from the Cambridge Structural Database (version
5.32, November 2010).
2 We use the term ‘‘neutral’’ arginine molecules to refer to those
cases (including zwitterionic forms) for which the overall charge on
the molecule is zero. In addition, the crystal structures of a number
of salts containing protonated arginine cations have been reported.
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