François Rieger

Electrophorus acetylcholinesterase: A glycoprotein; molecular weight of its subunits

The different forms of acetylcholinesterase (Ach E, EC 3.1 .1.7) from electric eel (Electrophorus electricus) have already been described, and characterized by their sedimentation coefficients in a sucrose gradient [ 1,2] . The physico-chemical characteristics of the three ‘native’ forms, A, C, D, with sedimentation coefficients 9.2 S, 14.2,S and 18.4 S, respectively, are only compatible with asymmetric structures. This hypothesis has now been confirmed by electron microscopy [3] . Another active, globular form G( 11.8 S) is obtained by tryptic, or other proteolytic treatment of A, C, and D; it appears as a tetramer in the micrographs. We have used the same preparations, that were studied by electron micrography, to investigate their subunit composition by SDS acrylamide gel electrophoresis. These results, coupled with those from electron microscopy, lead to specific proposals about the molecular structure of the various forms.

Phospholipids in ‘native’ Electrophorus acetylcholinesterase

Some time ago, our group showed that acetylcholinesterase, of Electrophorus electricus (electric eel) or Torpedo marmorata electric organs, exist in a number of molecular forms [ 11. The three original, or ‘native’ forms: A, C, and D, have the physico-chemical properties of asymmetric particles [2]. Globular forms can be derived from these native forms by tryptic treatment [ 1, 31, sonication [2] or through the action of electric eel endogeneous proteases [4, 51. A globular acetylcholinesterase, similar to that purified by other authors [6-81 can also be directly solubilized from the electroplax membrane by trypsin [9]. We have been able to obtain micrographs of both the globular and ‘native’ molecular forms we had described [lo]: these pictures clearly show the asymmetric, or ‘native’ forms, as a combination of a cluster of subunits, which we called the ‘head’, and an elongated element, which we called the ‘tail’. The active globular forms are tetramers (11.8 S) and dimers (7.7 S). We proposed a model in which the three native forms are composed of a tail and of, respectively, one, two, and three tetramers, for A (9 S), C (14.2 S) and D (18.5 S). We have indeed recently obtained micrographs of the D molecule, the ‘head’ of which is clearly made up of three clusters of four globules [I I]. Assuming a great stability of tetrameric association, it is thus possible to understand the occurrence of all five molecular forms, and some of their properties. The hydrodynamic properties of the native asymmetric forms of acetylcholinesterase in solution are consistent with the idea that these molecules behave like rigid structures of the dimensions observed in electron micrographs [ 121. Several authors have shown that acetylcholinesterase can be solubilized by proteases, but also

Interactions between lectins and electric EEL acetylcholinesterase

Multiple forms of acetylcholinesterase, solubilized from Electrophorus electricus electric organs have been characterized by their hydrodynamic properties and by electron microscopy observation [l-4] . They all have a higher CsCl isopycnic density (-1.31) than do the other proteins (-1.28), suggesting the presence of some sugar moiety. Polyacrylamide gel electrophoresis (with SDS and mercaptoethanol) of the purified enzyme has yielded two major protein sub-units, which positively stained with PAS reagent and contained sialic acid residues [5], The fact that neuraminidase has a pronounced action on the characteristics of the complement tixation of elongated forms of acetylcholinesterase [6] indicates that these sialic acid residues are part of some antigenic determinants of acetylcholinesterase. Furthermore, some results from Rosenberry et al. [7] suggest the presence of a certain amount of carbohydrate on the tetrameric purified form of acetylcholinesterase: amino acid analyses gave a content of 3-4% of hexosamines in this form. Ciliv and Ozand [8], working on erythrocyte acetylcholinesterase, made observations which also establish the presence of carbohydrates, in particular demonstrating that neuraminidase treatment changes electrophoretic mobility of acetylcholinesterase isozymes. Some more details have been given by Taylor et al. [9] on the presence of carbohydrates in Torpedo acetylcholinesterase: the total content of carbohydrates is 7.9% including hexoses, hexosamines and sialic acid, in a proteolysed 11 S form. This paper describes new evidence concerning the nature of the carbohydrate content of elongated forms of electric eel acetylcholinesterase, as suggested by the