Application to acetylcholine receptor

We have used the SEC/LALLS technique to determine the molecular weight of nicotinic acetylcholine receptor protein monomer, dimer and their subunits from the electric ray, Narke japonica [9, 11]. It is known that the receptor protein monomer is composed of four kinds of subunits, a, 6, Y and a. Fig. 5 A and B show, for example, elution profiles for the receptor monomer and the a-subunit, respectively. 

The molecular weights of the protein moiety of Narke acetylcholine receptor a, 3, Y subunits were determined to 39,000, 46,000, 47,000 and 48,000 respectively [11]. The values thus obtained are those for the protein moieties only, since (dn /dc)gjid/refers to the concentration of protein moieties In the subunits. Taking Into account the carbohydrate content of the subunits, the molecular weights were estimated to be 41,000, 49,000, 49,000 and 51,000 as listed in Table H. 

The molecular weights of the receptor protein moieties were also determined as above, but using 0.1 % sodium deoxycholate (NaDOC) as a surfactant Instead of SDS. The values obtained were 430,000 for the dimer and 220,000 for the monomer. Taking into account Che carbohydrate content (about 6 %) of Narke receptor proteins, the molecular weights were estimated to 460,000 for the dimer and 240,000 for the monomer. 

Considering that Narke receptor monomer has a subunit composition of (Ji28Ya [26], the protein moiety of the receptor monomer Is calculated to 219.000 from the values of the four kinds of subunits. This value agree well with the value 220,000 obtained for the protein moiety of the receptor monomer. 

The values of the molecular weights calculated from the primary structures of Torpede a, B, y and a subunits from the nucleotide sequences of cloned cDNA [27, 28] are about 20% higher than those obtained by us from SEC/LALLS. We cannot clearly explain the discrepancy. However, as pointed out by Noda et al. [27], it is likely that a carboxy-terminal peptide is eliminated by post-transitional cleavage. Furthermore, we cannot entirely eliminate the possibility that partial proteolytic digestions may occur in the process of preparation of the receptor proteins. 

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