R. G. Shulman

Transition Metal Binding in DNA Solutions

Paramagnetic ions in water shorten the spin‐lattice relaxation time T1 of the water protons. The effectiveness with which the relaxation takes place depends not only on the ion concentration, but also on the environment of the paramagnetic ion. In some cases where the paramagnetic ion is bound to a large molecule such as DNA, it shortens the proton relaxation time to a greater extent than when it is in solution. By measuring T1 by a pulsed nuclear magnetic resonance method, we have studied the binding of transition metal ions to DNA. We have obtained estimates for the number of available binding sites as well as information on the type of site which the various ions are bound to.

Luminescence of Pyrimidines, Purines, Nucleosides, and Nucleotides at 77°K. The Effect of Ionization and Tautomerization

The luminescence of purines and their corresponding nucleosides and nucleotides has been studied at 77°K for different states of ionization and for some alterations of tautomeric forms. To a first approximation, the excitation spectra always coincided with the absorption spectra. Furthermore the emission profiles were independent of the exciting wavelength and, except for the protonated forms of guanine and its derivatives, phosphorescence always consisted of a single exponential decay. The naturally occurring pyrimidines, uracil, thymine, and cytosine, fluoresced over a wide pH range but only phosphoresced at high pH where they have lost a proton. From a study of methylated pyrimidines it was shown that the lactim forms fluoresced at 300°K and phosphoresced at neutral pH at 77°K. In these two respects they differed from the natural pyrimidines and support the commonly accepted lactam structures for the natural pyrimidines.The purines, adenine and guanine, and their derivatives fluoresced and phosphoresce...

Proton Magnetic Resonance Shifts in Aqueous Solutions of Paramagnetic Metal Ions

The nuclear magnetic resonance of the water proton in aqueous solutions of paramagnetic metal ions is shifted by the isotropic hyperfine interaction AI·S with the electron spin of the metal. These shifts have been measured for solutions of Cr3+, Fe3+, Mn2+, Co2+, Ni2+, and Cu2+ between 0° and 100°C. Complete averaging of the shift due to rapid exchange of H2O in and out of the solvation shell occurred for Co2+, Cu2+, and Mn2+. However, the averaging of the shift was not complete for Cr3+, Fe3+ (and possibly Ni2+) and the kinetic parameters for the exchange process had to be considered in order to evaluate A.From the temperature dependence of the proton shifts in Cr3+ and Fe3+ solutions, we determined the pseudo first‐order rate constants k and the activation energies ΔE for the proton exchange between solvation shell and bulk. These values are: 8.7×104 sec−1 and 7.5 kcal mole−1 for Cr3+, 2.8×106 sec−1 and 12.1 kcal mole−1 for Fe3+. The proton exchange reaction proceeds via the acid dissociation of the hyd...

Molecular Orbital Analysis of Iron‐Group Cyanides

In the iron‐group cyanides the low‐lying antibonding orbitals of the cyanide ligands do not contain electrons. These π* orbitals play an important role in the covalency of the complexes. A molecular orbital analysis of their role is presented in order to correlate the orbital reduction factors derived from the ESR experiments of Baker, Bleaney, and Bowers with the isomer shifts of the Mossbauer experiments. By fitting the wavefunctions to the ESR experiment it is shown that the Fe2+ complex donates about one more electron to the π* orbitals than does the Fe3+. This shows that the effective charges of Fe2+ and Fe3+ are approximately the same and explains their similar isomer shifts.

Study of Metal‐Ion Binding to Nucleic Acids by 31P Nuclear Magnetic Resonance

We have measured the 31P NMR in aqueous solutions of RNA (ribonucleic acid) and AMP (adenosine monophosphate) containing Mn2+ and Co2+ ions. Measurements of T1, T2, and Δω, the shift from γIH0, were made as functions of the metal‐ion concentration and the temperature. Near‐room‐temperature linewidths were determined by the slow chemical exchange. At higher temperatures the Co2+ complexes enter the exchange‐narrowed region where 31P shifts in the CoAMP system are observed.The room‐temperature value of T1 for CoAMP and CoRNA complexes, the high‐temperature shifts of the CoAMP, and the high‐temperature values of T2 in the MnAMP all independently determine the strength of the isotropic transferred hyperfine coupling A between the metal ions and the 31P nuclei. Within the experimental accuracy which ranges from ∼5% (for Δω) to ∼50% (for T1 measurements) the values of A agreed with the values derived from 31P NMR measurements in crystals of Mn3(PO4)2 and Co3(PO4)2. From the similarity of A in crystals and compl...

BINDING OF MN2+ TO NUCLEIC ACIDS.

Measurements of the nuclear magnetic resonance relaxation times T1 and T2 of the water protons in aqueous solutions of nucleic acids and magnetic metal ions allow one to determine details about the metal‐ion binding. When Mn2+ is bound, the rotation of its hydration sphere becomes slower, so that its effectiveness in reducing the proton T1 from its value in pure water is enhanced. This relaxation enhancement has been measured for Mn2+ bound to DNA, RNA, synthetic polynucleotides, and E. coli ribosomes. Compared to water as unity, the enhancement factors range from 3.7 for polyuridylic acid to 16.7 for polyadenylic acid. The equilibrium constants for binding, and the concentration of binding sites have been calculated from the dependence of enhancement upon Mn2+ concentration.

Electron Spin Resonance of Irradiated DNA

The electron spin resonance (ESR) spectra of free radicals formed in-the sodium salt of desoxyribonucleic acid (DNA) by 1-Mev electrons were examined. Dry samples were irradiated at approximates 200°K and examined at 77°K at microwave power levels below approximates 0.1 mw. At levels below 1 Mrad only a single, rather broad absorption line was seen. At around this dosage, an additional spectrum containing several resolved hyperfine satellites was found. A free radical fragment compatible with the results was - C (CH 3)CH 2--. This free radical postulated must come from thymine which contains the only methyl group in DNA.

Electron Spin Resonance of the Excited Triplet States of Pyrimidines and Purines

The electron spin resonances of the metastable triplet states of various pyrimidines and purines have been observed in ethylene glycol: H2O glasses at 77°K. Generally the Δm=2 transitions were observed and the parameter of spin—spin interaction determined was (D2+3E2)½. However for orotic acid, guanylic acid, and adenylic acid it was possible to observe the Δm=1 transitions and to determine D and E separately. From the values of D and E and the decay times, which were all >0.2 sec, it was possible to characterize the lowest triplet states as π—π* states in all cases. For thymine, 5‐methylcytosine, and orotic acid the values of D (and E) agreed quite well with those calculated approximately for π—π* states from Huckel molecular orbitals.In addition to resonances attributed to neutral molecules, additional resonances were observed for the ionized states of azathymine, thymine, 5‐methylcytosine, and 5‐hydroxymethylcytosine. From the pH dependence of azathymine and thymine it was possible to show that one res...

Luminescence and Electron Spin Resonance Studies of Adenine in Various Polynucleotides

The excited states of adenine in different polynucleotides dissolved in ethylene glycol: H2O glasses have been studied by optical emission and ESR at 77°K. In single‐stranded poly A the optical properties were very similar to the constituent AMP monomers except for a red shift of the luminescence. The ESR Δm=1 transitions were observed with D=0.116 cm−1 and E=0.027 cm−1 for poly A and D=0.119 cm−1 and E=0.027 cm−1 (all ±0.002 cm−1) for AMP. This means that the triplet transfer time is longer than 2×10−10 sec since more rapid triplet—triplet jumping would give a time averaging of E. In the ordered double‐helix form of poly A the excited states are completely quenched when one‐half of the bases are protonated. In poly (A+2U) and poly (A+2I) ordered copolymers the excited states are also quenched. However in random distributions of U, C, and I in single strands of poly A no quenching was observed. This leads to the conclusion that the hydrogen bonding rather than energy transfer is responsible for the quench...

Measures of Covalency in Transition‐Metal Chlorides

The bonding between transition‐metal ions and chlorine is largely ionic. It has been known that the small degree of covalency in transition‐metal chloride crystals could be measured by transferred magnetic hyperfine interactions with the chlorine nuclei. Here we show that the Townes and Dailey theory allows one to derive the degree of covalency from the chlorine nuclear quadrupole coupling in these crystals. In the few cases where both magnetic hyperfine interactions and electric quadrupole interactions have been measured at the same nucleus, the degree of covalency calculated by the two methods agrees rather well. Both kinds of experiment indicate the importance of π bonding with the d orbitals as well as σ bonding.