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Structure determination: Nucleic acid
May 13, 2004
TIGP Chemical Biology and Molecular Biophysics
Lou-Sing Kan, Ph. D.
Institute of Chemistry
Structure and nomenclature of nucleic acid bases and sugars
Oligonucleotide chain (sugar-phosphate backbone)
The Watson-Crick base pairs The non Watson-Crick base pairs
The structures of DNA
CruciformSupercoil Tetra-stranded
Other structures of nucleic acids
Triple-stranded
Hairpin loop
Bulge loop
Internal loop
Mismatch
Knots
Nucleic acids: RNA and DNA
Ribonucleic Acids (RNA) composed by adenosine, guanosine, cytidine, uridine
Spectra of nucleosides in DMSO-d6
Adenosine
Cytidine
Guanosine
Uridine
www.aist.go.jp/RIODB/SDBS/menu-e.html
Deoxyribonucleic Acids (DNA) composed by deoxyadenosine, deoxyguanosine, deoxycytidine, thymidine
Comparison of nucleoside and deoxyribonucleoside in DMSO-d6
Comparison of nucleoside in DMSO-d6 and D2O
Adenosine
Deoxyadenosine
Cytidine in DMSO-d6
Cytidine in D2O
Structure and nomenclature of furanose and deoxyfuranose
Sugar-phosphate backbone conformation
The conformation of glycosidic bond
Proton NMR study on two structures of 3'-O-(Acetylimino)3'-de(phosphinico)-thymidylyl-(3’,5')-deoxythymidine in aqueous solution
O
O3'
NCAC
H3CO
OH
HONH
N
O
O
NH
N
O
O
'
NC
''
OAC
Tetrahedron 59 (2003) 1981-1988
Table I. Temperature dependent chemical shifts of TNT
TN H6 C5-CH3 H1 H2 H2 H3 H4 H5, H5 35℃ 7.48 1.74 6.13 2.47 2.21 4.69 4.14 3.63
25℃ 7.50 1.74 6.14 2.47 2.21 4.70 4.15 3.63
20℃ 7.50 1.74 6.14 2.46 2.21 4.71 4.15 3.63
15℃ 7.51 1.74 6.11 2.46 2.21 4.71 4.15 3.64
10℃ 7.51 1.73 6.12 2.49/2.41* 2.27/2.17 4.71 4.16/4.14 3.64
0℃ 7.53 1.73 6.14 2.49/2.41 2.27/2.17 4.73/4.71 4.17/4.13 3.64
NT H6 C5-CH3 H1 H2 H2 H3 H4 H5, H5 35℃ 7.29 1.74 6.09 2.24 2.32 4.34 4.02 3.87, 3.95
25℃ 7.30 1.74 6.10 2.24 2.32 4.33 4.03 3.88, 3.96
20℃ 7.31 1.74 6.11 2.23 2.32 4.32/4.39 4.03 3.88, 3.95
15℃ 7.32/7.28 1.74 6.11/6.19 2.23 2.32 4.32/4.40 4.04 3.90, 3.95
10℃ 7.33/7.28 1.73 6.12/6.21 2.21/2.21 2.31/2.36 4.32/4.41 4.04/4.01 3.90, 3.95
0℃ 7.34/7.29 1.73 6.14/6.23 2.21/2.21 2.31/2.36 4.33/4.42 4.06/4.01 3.94, 3.94
-COCH3
35℃ 2.03
25℃ 2.06/1.98
20℃ 2.07/1.96
15℃ 2.07/1.96
10℃ 2.07/1.95
0℃ 2.08/1.95
Table II. Coupling constants of sugar proton resonances
JH1-H2 JH1-H2 JH3-H2 JH3-H2 JH3-H4 JH4-H5 JH4-H5
TN 6.14 8.19 6.14 2.30 2.3 4.35 4.35
NT 6.66 6.66 6.14 6.14 4.10 7.42 3.58
major minor
N
3'T
O
O
CH3 5'T
N
3'T
O
H3C
5'TO
A) Conformation of major form of -5 C
B) Conformation of minor form of -5 C
1) 2)
2)1)
The Application of Chemical Shift
Proton magnetic resonance studies on the nonexchangeable protons of ribosyl ApApGpCpUpU
Biochemistry, 14 (1975) 4847-4863 & 4864-4869.
Biopolymers, 15 (1976) 2277-2286.
The Application of Coupling Constant
The Application of NOE
Detection of a guanine/adenine base pair in a decadeoxyribonucleotide by proton magnetic resonance spectroscopy
Proc. Natl. Acad. Sci., 80 (1983) 4263-4265.
Science, 238 (1987) 498-504.
(CCAAGATTGG)2
The Application of T1
Internal loop vs. hairpin
Biochemistry, 27 (1988) 3024-3031.
O
HO
N
N N
N
OH
NH2
H H
OH
2D COSY or DQF-COSY
2D TOCSY
2D NOESY
2D E.COSY
2D PE.COSY
Methods
ppm
2.53.03.54.04.55.05.56.06.5 ppm
2.5
3.0
3.5
4.0
4.5
5.0
5.5
6.0
6.5
7.0
Assignments based on structure
K. C. Wuthrich, NMR of Proteins and Nucleic Acids, John Wiley & Sons (1986)
The Study of Triplex Formation of TCTCTCCTCTCTAGAGAG
The assignment
Biophysical J., (2002) 82, 3170-3180.
References
Edwin D. BeckerHigh Resolution NMR, Theory and Chemical Applications, 3rd EditionAcademic Press, 2000.Ray FreemanMagnetic Resonance in Chemistry And MedicineOxford, 2003Joseph P. HornakThe Basic of NMRhttp://www.cis.rit.edu/htbooks/nmr/bnmr.htm
General
T.C. FarrarAn Introduction To Pulse NMR SpectroscopyFarragut Press, Chicago, 1987.H. Gunther"Modern pulse methods in high-resolution NMR spectroscopy."Angew. Chem.. Int. Ed. Engl.22:350-380 (1983)
Basic Pulse NMR
Ad BaxTwo-Dimensional Nuclear Magnetic Resonance in LiquidDelft University Press, 1982Richard R. Ernst, Geoffrey Bodenhausen, Alexander WokaunPriciples of NMR in One and Two DimensionsOxford, 1987
2D NMR
Peter BiglerNMR Spectroscopy Processing StrategiesVCH, 1997
Data Process
H. Duddeck, W. DietrichStructure Elucidation by Modern NMR, A WorkbookSpringer-Verlag, 1989
Application: small molecules
Kurt WuthrichNMR of Proteins and Nucleic AcidsJohn Wiley & Sons, 1986
C. A. G. HaasnootNMR in Conformation Analysis of Bio-organic Molecules
Application: Peptides and Proteins
Application: Nucleic Acids
G. C. K. RobertsNMR of Macromolecules, A Practical ApproachIRL Press, 1995.
Application: Others
S. Braun, H.-O. Kalinowske, S. Berger100 and More Basic NMR ExperimentsVCH, 1996S. W. HomansA Dictionary of Concepts in NMROxford, 1992Handbook of High Resolution Multinuclear NMRJohn Wiley & Sons, 1981
Dictionary