. The Biological bulletin. Biology; Zoology; Marine biology. -10 -8 -6 Log ([eserine]/M). -10 -8 -6 Log ([eserine]/M) Figure 1. Effect ofeserine (10"'^ Mto 10"' M) on acetylthiocholine hydrolysis in the soluble fractions of Eisenia felida (A) and Nereis div- ersicolor (B), Vertical bars denote SD of the mean for three determi- nations. buffers and sample preparation as described above for PAGE. Several methods for the histochemical detection of ChEs in polyacrylamide gels were assayed. All were based on the hydrolysis of a thiocholine ester and the formation of an insoluble copper co
. The Biological bulletin. Biology; Zoology; Marine biology. -10 -8 -6 Log ([eserine]/M). -10 -8 -6 Log ([eserine]/M) Figure 1. Effect ofeserine (10"'^ Mto 10"' M) on acetylthiocholine hydrolysis in the soluble fractions of Eisenia felida (A) and Nereis div- ersicolor (B), Vertical bars denote SD of the mean for three determi- nations. buffers and sample preparation as described above for PAGE. Several methods for the histochemical detection of ChEs in polyacrylamide gels were assayed. All were based on the hydrolysis of a thiocholine ester and the formation of an insoluble copper complex (Kamovsky and Roots, 1964; Koelle and Friedenwald, 1949). Original procedures were modified by elevation of the incubation pH to in Af phosphate buffer. ChE isoenzymes were identified through the use of a specific inhibitor of ChEs (10"^ Meserine sulfate) and by the comparative hy- drolysis of different thiocholine esters (ATCh, PrTCh, and BuTCh). Controls lacking substrate were prepared rou- tinely. Apparent molecular weights were measured with respect to the weights of standard proteins from Sigma (a-lactalbumin from bovine milk, 14,200; chicken egg al- bumin, 45,000; bovine serum albumin, 66,000 [mono- mer], 132,000 [dimer]; and jack bean urease, 272,000 [trimer], 545,000 [hexamer]). Results ChEs inhibition and substrate specificity The eserine sensitivity of ATCh splitting activity in a soluble (S) extract of A^. diversicolor or is shown in Figure 1. For E. fetida (Fig. lA) the curve is clearly biphasic, indicating the presence of at least two types of ChEs with different eserine sensitivities. The curve ob- tained for A', diversicolor (Fig. 1B) is monophasic, indi- cating the presence of only one kind of ChE. With 10"^ M iso-OMPA (inhibitor of pseudocholines- terases) and ATCh as substrate, extracts oi were inhibited by 20%, whereas no change was found with ex- tracts of N. diversicolor, suggesting that the ChEs of the latter speci
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