220 MHz Nuclear Magnetic Resonance Spectra of Oxidized and Reduced Pyridine Dinucleotides
Ramaswamy H. Sarma 1 and Nathan O. Kaplan 1
From the
1 From the Graduate Department of Biochemistry, Brandeis University, Waltham, Massachusetts 02154
1. The two C4 protons of the dihydropyridine ring of reduced coenzymes appear as an AB spectrum in the 220 MHz nuclear magnetic resonance system. This indicates that they are chemically and magnetically nonequivalent.
2. When the oxidized coenzyme is converted to the reduced coenzyme, the coupling constant of C'1H of ribose adjacent to the pyridine increased by about 3.5 Hz, whereas the same of the ribose adjacent to the adenine remained constant. This indicates that, when the oxidized pyridine dinucleotide is reduced, the ribose attached to the pyridine ring undergoes a conformational change, the conformation of the ribose attached to the adenine remaining the same in both oxidized and reduced coenzymes.
3. The C'1H of the ribose adjacent to pyridine of reduced coenzymes is substantially broadened, and this broadening increased at elevated temperatures. This is due to onset of unresolved nitrogen spin-spin coupling as a result of the alteration of the relaxation time of nitrogen dominated by quadrupole coupling. This observation provides the most unequivocal method for assigning the chemical shift of the C'1H of ribose attached to the pyridine of reduced coenzyme.
4. Startling changes take place in the shape and position of the ribose protons as a result of the reduction of the oxidized coenzyme. It is thought to result from an alteration in the geometric posture of the diphosphate backbone as a result of the reduction.
Submitted on November 4, 1968
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