G.E. Adams

Electron-affinic sensitization I. A structural basis for chemical radiosensitizers in bacteria

SummaryThe radiosensitizing properties of the highly electron-affinic nitroketone, para-nitroacetophenone (PNAP) have been investigated in resistant and sensitive bacteria, fern spores, mammalian cells in tissue culture and in suspensions of bacteriophage. Radiosensitization occurs in anoxic suspensions of Serratia marcescens Micrococcus radiodurans and in anoxic preparations of Chinese hamster cells line V79–379A. The sensitization of the mammalian cells (enhancement ratio 1·7 for 0·4 mM PNAP) is not inhibited in the presence of serum protein. No sensitization was observed for the spore of the fern Osmunda regalis.PNAP resembles oxygen in conferring some radioprotection on suspensions of bacteriophage T7 irradiated in broth.

Pulse radiolysis studies on the oxidation of organic radicals in aqueous solution

Pulse radiolysis has been used to measure directly the absolute rates of oxidation by ferricyanide ion of various radicals produced by OH attack on organic solutes. These include mono, di- and polyhydroxylic compounds, hydroxy acids, polyethylene oxides of molecular weight 200, 6000 and 20 000 and the amino acid serine. Radicals produced by hydrogen abstraction from α carbon atoms in alcohols are oxidized at, or near, diffusion-controlled rates, whereas the reactions are much slower for radicals formed by OH-attack elsewhere. The technique has been used to measure the percentage OH-attack at the α position for a series of straight and branched-chain alcohols. n Oxygen competes with ferricyanide for radical oxidation. The data for oxygen-containing solutions fit a simple radical-competition scheme which has been used to measure rates of peroxy-radical formation. These approach diffusion-controlled limits.

Electron Affinic Sensitization: Part II: Para-Nitroacetophenone: A Radiosensitizer for Anoxic Bacterial and Mammalian Cells

SummaryThe radiosensitizing properties of the highly electron-affinic nitroketone, para-nitroacetophenone (PNAP) have been investigated in resistant and sensitive bacteria, fern spores, mammalian cells in tissue culture and in suspensions of bacteriophage. Radiosensitization occurs in anoxic suspensions of Serratia marcescens Micrococcus radiodurans and in anoxic preparations of Chinese hamster cells line V79–379A. The sensitization of the mammalian cells (enhancement ratio 1·7 for 0·4 mM PNAP) is not inhibited in the presence of serum protein. No sensitization was observed for the spore of the fern Osmunda regalis.PNAP resembles oxygen in conferring some radioprotection on suspensions of bacteriophage T7 irradiated in broth.

Selective Free Radical Reactions with Proteins and Enzymes: The Inactivation of Ribonuclease

ADAMS, G. E., BISBY, R. H., CUNDALL, R. B., REDPATH, J. L., AND WILLSON, R. L. Selective Free Radical Reactions with Proteins and Enzymes: The Inactivation of Ribonuclease. Radiat. Res. 49, 290-299 (1972). The effect of free radicals derived from thiocyanate, bromide, and carbonate ions on the activity of ribonuclease have been studied in neutral and alkaline solutions. Thiocyanate ions and, to a lesser extent, carbonate ions, protect ribonuclease against radiation-induced inactivation, although the presence of bromide ions has little effect. The absolute rate constants for the reactions of these radical anions with ribonuclease have been measured as a function of pH and salt concentration, and the transient absorption spectra of the reaction products have been determined. The results suggest that damage to a histidine residue, or residues, leads directly to loss of activity and support previous conclusions that exposed tyrosine residues in the enzyme are not essential for activity.

On the Mechanism of the Radiation-induced Inactivation of Ribonuclease in Dilute Aqueous Solution

SummaryPulse radiolysis data from ribonuclease solutions have been used to aid the interpretation of the mechanism of the radiation-induced inactivation of this enzyme. Activity measurements show that in γ-irradiated dilute aqueous solution, hydrated electrons do not contribute to the inactivation process. In deoxygenated solution, 90 per cent of the inactivation process is due to reactions of OH radicals with the remainder attributable to reactions of hydrogen atoms.Pulse radiolysis studies show the formation of three transient spectra which have been assigned respectively to reactions of OH, H and eaq− with the enzyme. It is concluded that the spectrum of the OH-induced transient contains contributions from OH-adduct spectra of the ring-containing amino acids and that some of the short-lived species are involved in the initial stages of the inactivation process.

Pulse radiolysis of sulphur compounds. Part 3.—Repair by hydrogen transfer of a macromolecule irradiated in aqueous solution

Free radical repair by hydrogen transfer from a sulphydryl compound (cysteamine) has been extended to a polymeric system. Repair rate constants vary from 2.6 × 107 M–1 sec–1 for ethylene glycol to 5 × 106 M–1 sec–1 for polyethylene oxide (PEO) of molecular weight 20,000. The analysis of rate data for polymers of m.w. >200 is complicated by competing radical-radical reactions. A comprehensive reaction scheme is proposed and the kinetic equations solved numerically. The effects of pH and molecular weight on the repair rate have also been studied. Some further data on the pulse radiolysis of cysteamine solutions, including a determination of the extinction coefficient of the radical-ion RSSR–(Iµ4100= 9.4 ± 0.5 × 103 M–1 cm–1) are presented.

Radiation effects on

ADAMS, G. E., BAVERSTOCK, K. F., CUNDALL, R. B., AND REDPATH, J. L. Radiation Effects on a-Chymotrypsin in Aqueous Solution: Pulse Radiolysis and Inactivation Studies, Radiat. Res. 54, 375-387, (1973). Pulse radiolysis and steady-state inactivation methods have been used to study the separate roles of H, OH, and e.q- in the radiation-induced inactivation of aqueous a-chymotrypsin solutions. The effects of nitrous oxide and t-butanol used as selective free radical scavengers allow the inactivating efficiencies per unit G-value of OH, H, and eaq- radicals to be estimated; these are 0.137, 0.13, and 0.055, respectively, in neutral solution. The transient spectra observed show that both OH radicals and hydrogen atoms react mainly with tryptophan residues. Only about 15% of the hydrated electrons react with the cystine disulfide linkages; rupture of some of these linkages may lead to inactivation. It is suggested that the protective action of oxygen is due partly to the reactions of 02 with enzyme radicals formed initially by reaction with OH.

alpha

SummaryThe effect of the selective free radicals (CNS)2− and Br2− on the activity of papain has been studied in neutral and alkaline solution. These radicals are more efficient than the OH radical for inactivating papain in neutral solution. In alkaline solution, OH and Br2− are equally effective, whereas (CNS)2− is less efficient than OH.The absolute rate constants for the reactions of these radicals with papain have been measured as a function of pH for both active and inactive preparations of the enzyme. The transient absorption spectra of the reaction products have also been determined.The results show that tryptophan residues as well as cysteine-25 are essential to the activity of papain.

-chymotrypsin in aqueous solution: pulse radiolysis and inactivation studies

SummaryThe reactions of the selective free radicals (CNS)2− and Br2− with α-chymotrypsin have been studied by pulse radiolysis and steady-state inactivation measurements.Rate-constants for the reaction have been measured over the pH range 7 to 12. The reactivity increases at pH 11 where the tyrosine residues deprotonate. Transient product spectra also indicate a significant increase in attack at tyrosine at this pH. A small increase in rate was observed by lowering the salt concentration from 10−1 M to 4 × 10−2 M.The inactivation data show (CNS)2− to be protective relative to OH over the pH range 2 to 11, whereas Br2− sensitizes between the pH range 4 and 9.From the pulse radiolysis and inactivation data, it was concluded that a histidine residue was essential to the activity of α-chymotrypsin.

Selective Free-radical Reactions with Proteins and Enzymes: Pulse Radiolysis and Inactivation Studies on Papain

The enzyme D-amino acid oxidase and its apoenzyme have been irradiated at pH 5.5--10 under conditions designed to assess the inactivating effect of OH radicals and the selective free radicals Br2- and (SCN)2-. Near neutral pH, removal of the coenzyme FAD from the enzyme results in greater inactivation by selective free-radical attack. From pulse-radiolysis spectra, this increase is associated with attack on tyrosine and tryptophan residues in the protein. A large increase in inactivation of both the haloenzyme and apoenzyme by selective free-radical attack is seen with increasing alkalinity. This is consistent with attack on tyrosine being of major importance.