X-ray structural determination for 5'-CMP·Na2 mononucleotide
Gh. Mihailescu1, Al. Darabont2, Gh. Borodi1, I. Bratu1, Mihaela Pop2
1
National Institute for Research and Development of Isotopic and Molecular
Technology, P.O. Box 700, R-3400 Cluj-Napoca 5, Romania
2
”Babes-Bolyai” University, Dept. of Physics, Cluj-Napoca, Romania
Abstract
The paper presents the results obtained in the growing and structure determination by X-ray diffraction of the
5'-CMP.Na2 mononucleotide (5'-cytidine monophosphate disodium salt) single crystal, component of the ARN.
The space group, the unit cell parameters and the fractional coordinates were determined.
Introduction
Cytidine 5’-monophosphate disodium salt (5’-CMP.Na2) is one of the four common
ribonucleotides which make up ribonucleic acids (RNA’s). The crystal structure of the sodium
salt of cytidine 5’-monophosphate 6.5 hydrate provides valuable information on the
interaction mechanisms of alkaline metal ions with water and nucleotides. Previous studies
have been done on the barium salt of cytidine 5’-monophosphate 8.5 hydrate [1] and some
information on the crystal data of disodium cytidine 5’-monophosphate were indicated by
these authors. Nevertheless, the molecular structure and conformational parameters are not
published yet to our knowledge. Nishimura et al. [2] indicated that the crystal structure of this
salt was not solved. Difficulties in growing single crystals of this mononucleotide would be
the reason. Despite these difficulties single crystals have been obtained and the corresponding
crystal structure has been determined in this work.
The nucleosides and nucleotides have attracted considerable attention not only because
they are building blocks of the nucleic acids, DNA and RNA, which are the center of life
processes, but also because they are cofactors and allosteric effectors for many of the
fundamental enzymatic reactions. Cytidine 5’-monophosphate disodium salt (5’-CMP.Na2) is
one of the four common ribonucleotides which make up ribonucleic acids (RNA’s).
Results and Discussion
The 5’-CMP Na2 compound was crystallized by the slow evaporation method from
water-methanol solution using three recrystallization processes. There were obtained
transparent single crystals of about 2 x 4 x 1-2 mm. From the Laue method we put in evidence
the presence of the 2-fold symmetry. By Kulpe method the crystal was oriented along the 2fold axis and by the oscillation method it was determined the periodicity along this direction,
b=8.899Å. The spot-layer of superior order proved to be symmetrically disposed relative to
the spot layer of zero order indicating again the 2 order symmetry. In the Fig. 1 the oscillation
pattern is shown.
Fig 1: The oscillation pattern of the 5'-CMPNa2 . 6.5 H2O single crystall
From the zero layer Weissenberg pattern [3], Fig. 2, the reciprocal lattice parameters resulted
as: a*=0.137, c*=0.122 and *=85.5. It results a=14.08 Å, c=15.83 Å and =94.5.
Fig. 2: The zero layer Weisenberg pattern for the 5'-CMPNa2 . 6.5 H2O single crystall
By indexing the powder diffraction pattern there were obtained more precise the lattice
parameters. The compound density was experimentally determined by the picnometric
method, exp = 1.999 g/cm3. From the lattice parameters and the experimental density resulted
the molecules number in the unit cell: Z = 4.
Therefore, the 5’-CMP.Na2 compound crystallizes in the monoclinic system, the space
group P21 with 4 molecules per unit cell and the lattice parameters are: a = 14.10Å, b = 8.9Å c
= 16.05 Å and  = 94.
Intensity data up to sin/=0.594Å-1 were measured on a Nonius CAD4 diffractometer
with MoK (0.71069 Å) radiation in the -2 scan mode from a 0.3 x 0.35 x 0.35 mm single
colorless crystal of prismatic shape. A number of 3756 unique reflections (Rint = 0.094) were
collected out of which 2202 are greater than 2(I). The (-7 0 1) and (-2 1 6) reflections were
monitored during data collection to check crystal and instrument stability and no appreciable
change in their intensities was detected. The unit cell parameters were refined by least-square
refinement of the 2 values of 25 strong well centered reflections in the range 13<2<26.
The refined unit cell dimensions are a = 14.042(5), b = 8.924(5), c = 16.091(5) Å,
 = 94.410(5) , Z = 4, the cell volume is V = 2010.4 Å3.
The structure was solved by direct methods with SIR97 [4] and SHELX97 [5] software.
Almost all the atom positions in the structure were obtained from an E map computed for the
best set of phases. Some oxygen atoms from water molecules were subsequently located from
difference Fourier maps. The ulterior least-square refinements with anisotropic temperature
factors for all the atoms converged with R = 0.065 for the 2202 reflections exceeding 2(I).
The H atoms of the nucleotide molecules and three O atoms from water molecules were
automatically fixed and isotropically refined. The other ten water molecules in the unit cell
were found to be disordered, generating alternative solvent layers between the nucleotide
molecules. The final parameters for the non-H atoms and for the O atoms of the water
molecules are listed in Tables 1 and 2, respectively.
Conclusions
The 5'-CMP.Na2 single crystall was grown from a mixture of water-methanol solution and
crystallyzes in the monoclinic crystallographic system with four molecules per unit cell and
the space group P21. The unit cell parameters and the fractional coordiantions were
determined.
References
[1] J. Hogle, M. Sundaralingam and J.H.Y. Lin, Acta Cryst. 1980, B36, 564-570.
[2] Y. Nishimura, M. Tsuboi, T. Sato, and K. Aoki, J.Mol.Struct., 1986, 146, 123-153.
[3] G.H. Stouth and L.H. Jensen, "X-Ray Structure Determination, a Practical Guide",
2nd Edition, The Macmillan Comp., New York, 1989
[4] A. Altomare, G. Cascarano, C. Giaccovazzo, A. Guagliardi, A.G.G. Moliterni, M.C.
Burla, G. Polidori, M. Camalli, R. Spagna, SIR97, A package for crystal structure solution by
direct methods and refinement, 1997, Univ. of Bari, Italy.
[5] G. M. Sheldrick, SHELX 97, Program for crystal structure determination, Univ. of
Heidelberg, 1997, Göttingen, Germany.
[5] W. Saenger, Principles of Nucleic Acid Structure, 1984, Springer Verlag, New York,
Berlin, pp. 15-24
[6] Landolt-Börnstein, Vol. VII: Biophysics, Vol. 1 “Crystallographic and Structural Data”,
ed. W. Saenger, Springer Verlag, Berlin, Heidelberg, 1989, pp. 6-21.
[7] Ch. L Barnes, S.W.Hankinson, Acta Cryst., 1982, B38, 812-817.
[8] S.K. Kattl, T.P. Seshadri, M.A. Viswamitra, Acta Cryst. 1981, B37, 1825-1831.
Table 1. Fractional coordinates (Å) and isotropic equivalent temperature factor (Å2) with s.u.
in parentheses
Molecule A
Molecule B
X
Y
Z
Ueq
X
Y
Z
Ueq
0.16982(19)
-0.0821(4)
-0.06485(17)
0.0234(7)
0.3933(19)
0.1951(3)
0.63979(17)
0.0203(7)
N(1)
0.8618(6)
0.5176(12)
0.7307(6)
0.030(2)
0.7820(6)
0.0938(11)
0.6595(5)
0.026(2)
C(2)
0.8592(8)
0.5590(15)
0.6479(7)
0.026(3)
0.8767(7)
0.0448(15)
0.6698(7)
0.027(3)
O(2)
0.9275(6)
0.5240(12)
0.6059(5)
0.041(2)
0.9296(5)
0.0802(12)
0.6147(5)
0.039(2)
N(3)
0.7854(7)
-0.3643(13)
0.6134(6)
0.032(2)
0.9084(7)
-0.0284(12)
0.7380(6)
0.030(2)
C(4)
0.7112(7)
-0.3303(13)
0.6578(7)
0.025(3)
0.8471(7)
-0.0594(14)
0.7956(7)
0.025(3)
N(4)
0.3614(7)
0.2385(12)
0.3806(6)
0.034(3)
0.8829(7)
-0.1280(12)
0.8651(6)
0.031(2)
C(5)
0.7117(8)
-0.3676(15)
0.7426(7)
0.030(3)
0.7498(8)
-0.0162(15)
0.7862(7)
0.029(3)
C(6)
0.7864(8)
-0.4474(15)
0.7759(7)
0.031(3)
0.7199(8)
0.0563(15)
0.7172(7)
0.026(3)
C(1’)
0.0604(8)
-0.0741(15)
0.2310(6)
0.027(3)
0.7490(7)
0.1850(14)
0.5880(6)
0.023(2)
C(2’)
-0.0009(8)
-0.4942(15)
-0.1611(7)
0.030(3)
0.6783(8)
0.1108(14)
0.5267(7)
0.024(2)
O(2’)
0.0690(6)
-0.4029(10)
-0.1922(5)
0.036(2)
0.7239(5)
0.0247(9)
0.4687(5)
0.033(2)
C(3’)
-0.0377(8)
-0.1289(15)
0.1098(7)
0.030(3)
0.6272(8)
0.2486(13)
0.4889(6)
0.024(2)
O(3’)
-0.1214(6)
-0.1793(12)
0.1451(6)
0.044(2)
0.6849(6)
0.3042(11)
0.4269(5)
0.039(2)
C(4’)
0.0439(8)
-0.2426(15)
0.1207(7)
0.029(3)
0.6269(7)
0.3577(14)
0.5593(6)
0.023(2)
O(4’)
0.1006(6)
-0.2012(10)
0.1960(5)
0.0319(19)
0.7020(5)
0.3088(9)
0.6203(5)
0.0288(18)
C(5’)
0.1082(9)
-0.2512(16)
0.0504(7)
0.035(3)
0.5349(8)
0.3644(14)
0.6035(8)
0.030(3)
O(5’)
0.1373(6)
-0.1051(9)
0.0283(5)
0.0307(19)
0.5035(5)
0.2165(9)
0.6213(5)
0.030(2)
O(I)
0.0880(5)
-0.1335(9)
-0.1250(5)
0.0294(19)
0.3352(6)
0.2475(10)
0.5632(5)
0.033(2)
O(II)
0.1899(6)
0.0814(9)
-0.0704(5)
0.032(2)
0.3748(6) 0.2886(10)
0.7161(5)
0.032(2)
O(III)
0.2579(5)
-0.1760(10)
-0.0739(5)
0.0298(19)
0.3838(5)
0.0297(9)
0.6582(5)
0.0293(19)
Na(1)
0.9163(3)
0.4168(7)
0.2672(3)
0.0427(13)
0.2778(10)
0.279(2)
0.0353(9)
0.216(9)
Na(2)
0.7664(3)
0.1465(6)
0.3426(3)
0.0404(13)
0.1729(4)
0.1898(10)
0.5252(3)
0.069(2)
P
Table 2. Fractional coordinates (Å) and isotropic equivalent temperature factor (Å2) with s.u.
in parentheses for the oxygen atoms of the water molecules.
X
Y
Z
Ueq
O(W1)
0.7467(5)
0.3619(11)
0.2497(5)
0.040(2)
O(W2)
0.8063(6)
-0.0992(12)
0.2994(6)
0.047(2)
O(W3)
0.9229(6)
0.2336(12)
0.3739(6)
0.046(3)
O(W4)
0.6046(8)
0.0695(12)
0.2315(7)
0.065(3)
O(W5)
0.3753(6)
0.1765(12)
0.8755(5)
0.044(2)
O(W6)
0.4992(6)
-0.1763(12)
0.7343(5)
0.043(2)
O(W7)
0.2782(9)
-0.4706(13)
-0.0972(7)
0.066(3)
O(W8)
0.8874(7)
0.6273(13)
0.3479(6)
0.056(3)
O(W9)
0.5325(8)
-0.0234(15)
0.8900(6)
0.065(3)
O(W10)
0.4105(8)
-0.2099(17)
0.0388(8)
0.081(4)
0.073(2)
0.0975(9)
0.115(6)
O(W11) 0.4536(12)
O(W12)
0.9568(7)
-0.13711(7)
0.4958(6)
0.057(3)
O(W13)
0.4228(6)
0.3294(19)
0.9945(6)
0.106(7)