Radiolytic reduction of protein and nonprotein disulfides in the presence of formate: a chain reaction.
We have reported recently that the disulfide groups in bovine serum albumin can be reduced by a radiolytic chain reaction which occurs in deoxygenated solutions containing formate ions. This reaction, which involves the reduction of disulfide groups by hydrated electrons and carbon dioxide radical anions, has now been studied in greater detail and compared with an analogous reaction in small, disulfide containing molecules over a range of pH values and substrate concentrations. A two-step reaction is proposed to account for the reduction of disulfides in reactions which can have chain lengths of 20 or more. Thiols produced by the disulfide reduction are stable to the conditions of the reaction. For example, a biological assay showed that the integrity of glutathione was maintained even at radiation doses much larger than those required to achieve complete reduction of glutathione disulfide. It was found that the extent of disulfide reduction could easily be controlled by varying the radiation dose delivered to the solutions. Radiolytic reduction is a very useful way of reducing protein and low molecular weight disulfides without the use of excess quantities of reagents such as dithiothreitol. In many cases, the reaction solutions could be used directly for subsequent reactions and this may be of considerable value in modifying the structure of hormones, enzymes, membrane receptors, and other disulfide containing proteins. If ammonium formate is used, freeze drying is an effective way to remove the formate salt, should this be required.
Koch
C J
CJ
University of Pennsylvania, Philadelphia 19104-6072.
Raleigh
J A
JA
D002588:Cesium Radioisotopes; D003037:Cobalt Radioisotopes; D004220:Disulfides / Q000737:chemistry* / Q000528:radiation effects; D005561:Formates / Q000737:chemistry*; D010084:Oxidation-Reduction; D011506:Proteins / Q000737:chemistry* / Q000528:radiation effects; D012710:Serum Albumin, Bovine / Q000378:metabolism / Q000528:radiation effects
D002588:Cesium Radioisotopes; D003037:Cobalt Radioisotopes; D004220:Disulfides; D005561:Formates; D011506:Proteins; C030544:formic acid; D012710:Serum Albumin, Bovine