REACTIONS WITH PROTEINS 757 



served by Riggs (Gibson and Houghton, 1955). Oxygenation of hemoglobin 

 facilitates reaction of the SH groups with A^-ethylmaleimide and iodoaceta- 

 mide but not with p-MB (Benesch and Benesch, 1962). The mechanisms 

 by which these effects are produced are not clear, but hypotheses have been 

 offered based on the spatial arrangement of the SH groups and the hemes. 

 Riggs (1959) considers the hemoglobin molecule to consist of two halves, 

 each with a pair of reactive SH groups and a pair of hemes, the SH groups 

 perhaps lying between the hemes. Any mercurial which can form a bridge 

 between the 2 SH groups of a pair — such as Hg++, or mersalyl if the 

 C — Hg bond is ruptured and inorganic Hg++ is released — increases the 

 affinity of the hemes for Og, whereas mercurials reacting only with a single 

 SH group — such as p-MB and MM — do not have this effect. Any interac- 

 tion with the SH groups reduces the interaction between the hemes, prob- 

 ably by bringing about reversible structural changes in the protein config- 

 uration. On the other hand, Klotz and Klotz (1959) favor a mechanism 

 involving disturbances in the water structure around and between the 

 hemes. Whatever the explanation, the bearing on the effects of mercurials 

 on enzyme active centers by reaction with adjacent SH groups is obvious. 



(C) Mercaptalbumin. Mercaptalbumin is one fraction of the serum al- 

 bumins containing a single reactive SH group whereas the other albumins 

 contain none. It was isolated as the crystalline mercury salt by Hughes 

 (1947) and its reactions have been studied in detail, so that it has become 

 the classic example of protein-mercurial interaction (W. L. Hughes, 1950). 

 The three major reactions maj^ be represented as follows: 



Reaction 1: alb— SH + RgX^ ±5 alb— S— HgX + H+ 4- X" 



Reaction 2: alb— S— HgX + alb— SH ±^ alb— S— Hg— S— alb -f- H+ -^ X- 



Reaction 3: alb— S— Hg— S— alb + RgX^ ^ 2 alb— S— HgX 



where alb indicates mercaptalbumin and X some ligand (e.g. Cl~). The 

 first reaction is mercaptide formation, the second dimerization, and the 

 third dissociation of the dimer by excess HgXg. Formation of the dimer 

 increases the turbidity and, if some ethanol is added, crystals form. These 

 crystals are colorless diamond-shaped orthorhombic plates, containing chan- 

 nels or enclosures of fair size, with liquid within them, and permeable to 

 various salts, sugars, and dyes (Low and Weichel, 1951). Hughes and Dint- 

 zis (1964) have described procedures for crystallizing the dimers from etha- 

 nol-water mixtures at low temperatures. Viscosity and sedimentation stud- 

 ies (Low 1952) led to the representation of the dimer as in A of Fig. 7-5, 

 while the results of X-ray diffraction study are compatible also with struc- 

 ture B. The lengths of the dimer would be around 140-150 A, monomer 

 mercaptalbumin being of molecular weight 66,000. The structure is inde- 

 pendent of the smaller ions making up the crystal; e.g., the dimer will 



