A PROPOSED MECHANISM OF PROTEIN 

 IN ACTIVATION* 



L. G. AUGENSTINE 

 Brookhaven National Laboratory, Upton, Long Island, New York 



Abstract— An hypothesis dealing with the role of disulfide bonds in protein inactivation by 

 physical agents has been discussed with reference to material presented at this conference. 

 It is proposed that the critical effect becomes localized at a 'weak-link', causing first the 

 rupture of a disulfide bond, followed by the breaking of neighboring intramolecular bonds 

 and finally the rupture of a second disulfide bond. Much of the evidence upon which these 

 postulates are based is reviewed. The manner in which this model defines a target volume is 

 indicated and alternative methods of disulfide splitting are discussed. 



The author has previously proposed (I) an hypothesis deahng with the general 

 problems of protein inactivation and the importance of disulfide bonds in 

 maintaining protein structure. This hypothesis was originally presented to 

 account for heat denaturation data. It has since been extended in an attempt 

 to account for inactivation by ultraviolet hght and by ionizing radiation 

 ('direct effect') (2, 3, 4). The model is a special case of more general ones 

 proposed by Mirsky and Pauling (5), Lumry and Eyring (6) and Platzman 

 and Franck (7), and would depend for its accomphshment upon physical 

 processes similar to those described by the latter authors. It is to be emphasized 

 that this scheme is not advanced as the only mechanism whereby protein 

 inactivation can occur, but rather as the most likely. 



It is proposed that the critical effect of the physical agents mentioned is 

 not to cause indiscriminate molecular disorganization. Rather, their primary 

 effect becomes preferentially locaHzed at certain points in the molecule! . 

 Further, certain of these points (collectively called the 'weak-hnk') are involved 

 in processes which are characteristic of all proteins and which lead to inactivation. 

 These processes can be characterized as occurring in three distinct steps. 



1. The breaking of an S — S bond; 



2. The breaking of a variable number of neighboring intramolecular bonds 



(e.g. H-bonds); and 



3. The rupture of a second S — S bond. 



Step 1 requires about 20 kcal/mole, or 0.9 eV per molecule (8), and a 

 negligible entropy factor, while an appreciable entropy increase is associated 

 with step 2. Step 3 allows the spontaneous formation of a structure incom- 

 patible with further activity. Although irreversibility could result from the 



* Research carried out at Brookhaven National Laboratory under the auspices of the U.S. 

 Atomic Energy Commission. 



t For instance, Platzman (4, p. 19) has pointed out that 'the most stable position for a 

 migrating electron vacancy to become localized is at a site that can be crudely identified with the 

 atom of lowest ionization potential'. 



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