Precise Functional Analysis of Biological Macromolecules Promoted by the Encounter of Crystallographic and Spectroscopic Molecular Structure Analyses Takamitsu Kohzuma*, Sachiko Yanagisawa, and Tomotake Niizeki Faculty of Science, Ibaraki University The three dimensional structure of protein molecule is an essential for the comprehension of numerous life phenomena and the industrial utilization of biological systems. It is important to determine the three dimensional structure of biological macromolecule components by a certain techniques, X-ray crystallographic structure analysis, neutron crystallographic structure analysis or NMR. Although these structure analyses are very sophisticated methods for the determination of the biological macromolecule, the spectroscopic techniques are very important for the complete understanding of the time-dependent biological phenomena accompanying with the structural alternation of the relevant functional components, such as proteins and nucleic acids. The UV resonance Raman spectroscopy is introduced as an example for the precise local structure analysis with the X-ray crystallographic structure of protein molecules. DNA X NMR X X X
H + Vol. 7 (No. 2), NMR
1000 O 2 O 2 O 2 T R T R T 215cm -1 R 221cm -1 T R T R Kitagawa T O 2 H + C 13 =C 14 K L M N O / Cu(II) Dave S Cu(II)-S - Cu-S - Kohzuma
EPR S 195 nm 210 nm 206.5 nm Kr + 413.1 nm 280 nm 235 nm 470 244 nm Ar + 488 nm 260 nm 260 nm 257 nm
(a) ph 4.0 (b) ph 6.9 (c) ph 11.0 (a) 244 nm 244 nm (b) 244 nm (a)
ph (b) (c) 244 nm 1616 cm -1 (Y8a) 1604 cm -1 1600 cm -1 pk a 880 cm -1 W17 NH 1360 / 1340 cm -1 I (1360) / I (1340) 1 1 C α C β -C 3 C 2 2 χ 2,1 11 NCN ND 12 N ph* pk a 1602 cm -1 C 4 C 5 C α C β -C 4 C 5 χ 2,1 13 1385 cm -1 1350 cm -1 1602-1 cm C 4 C 5 1580 1560 cm -1 14 10 8
T R ph 229nm Bohr 15 Nagai His 87 16 (HRP) IX 6 His42( His) Arg38 Asn70 2.0 X Hashimoto 17 SOD O - 2 O 2 H 2 O 2 SOD O OD 18 235 nm 19 14000 124 600 nm Cys78 Met86 His40 His81 20
244 nm H 2 O D 2 O H81 M86 Cu C78 H40 (a) (b) Cu(II)PAz 235 nm Cu(II)PAz 1612 cm -1 1206 cm -1 1176 cm -1 1469 cm -1 1448 cm -1
1384 cm -1 His6 1404 cm -1 1283 cm -1 (Cu(I)PAz) 1384 cm -1 1408 cm -1 His81 244 nm Cu(II)PAz 1384 cm -1 Cu(I)PAz 15 cm -1 Cu(I)PAz 21 (II) (II) ( ) Cu(II)PAz Cu(I)PAz
(235 nm (235 nm ) ) (235 nm ) (235 nm )
Photosystem I Photosystem II 22 Photosystem I Freeman 23 Freeman
24 Sykes 25 acidic patch acidic patch Ubbink 26 Hirota 27 597 nm 590 nm 387 mv 20 mv 28 28 29 Yamauchi 30 acidic patch
31 Kostic
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