Protein gamma-radiolysis in frozen solutions is a macromolecular surface phenomenon: fragmentation of lysozyme, citrate synthase and alpha-lactalbumin in native or denatured states.
OBJECTIVE
To test whether radiolysis-induced fragmentation in frozen aqueous protein solution is dependent on solvent access to the surface of the protein or to the molecular mass of the polypeptide chain.


METHODS
60Co gamma-irradiation of three proteins at -78 degrees C: lysozyme, citrate synthase and alpha-lactalbumin in their native state, with or without bound substrate, or denatured (random coil in urea/acid-denatured state).


RESULTS
By SDS-polyacrylamide gel electrophoresis/analysis of the protein-fragmentation process, it was found that for a given protein D37 values (dose to decrease the measured amount of protein, with an unaltered polypeptidic chain, to 37% of the initial amount) varied according to the state of the protein. D37 for denatured proteins was always much smaller than for native states, indicating a greater susceptibility to fragmentation. In urea, contrary to the native state, no well-defined fragments were observed. Radiolysis decay constants (K= 1/D37) increased with solvent-accessible surface area of these proteins estimated from their radii of gyration in the various states. This is shown also in previous data on native or SDS-denatured proteins. Denatured proteins which have a large surface area exposed to the solvent compared with native ones are more fragmented at equal doses.


CONCLUSIONS
It is concluded that D37 is directly related to the exposed surface area and not to the molecular mass of the polypeptide chain.
Audette
M
M
LPCR, UMR 8610 CNRS-Université Paris-Sud, Orsay, France.
Chen
X
X
Houée-Levin
C
C
Potier
M
M
Le Maire
M
M
D000818:Animals; D002645:Chickens; D002950:Citrate (si)-Synthase / Q000378:metabolism*; D003037:Cobalt Radioisotopes; D004591:Electrophoresis, Polyacrylamide Gel; D005615:Freezing; D005720:Gamma Rays; D006168:Guinea Pigs; D007768:Lactalbumin / Q000378:metabolism*; D008024:Ligands; D008956:Models, Chemical; D015233:Models, Statistical; D009113:Muramidase / Q000378:metabolism*; D009206:Myocardium / Q000201:enzymology; D011487:Protein Conformation / Q000528:radiation effects; D011489:Protein Denaturation / Q000528:radiation effects; D017510:Protein Folding; D013552:Swine; D014508:Urea / Q000378:metabolism
D003037:Cobalt Radioisotopes; D008024:Ligands; D014508:Urea; D007768:Lactalbumin; D002950:Citrate (si)-Synthase; D009113:Muramidase