BIOCHEMICAL AND MOLECULAR CHARACTERIZATION OF ACETYLCHOLINESTERASE FROM THE HAGFISH MYXINE GLUTINOSA

To obtain information about the evolution of the cholinesterases, we i nvestigated the cholinesterase activity of an agnathan vertebrate, the hagfish Myxine glutinosa. On the basis of evidence from enzymology, p harmacology, and molecular biology, we conclude that the cholinesteras e activity is due to acetylcholinesterase (AChE). The enzyme hydrolyze s acetylthiocholine preferentially and exhibits substrate inhibition. The hydrolysis of both acetylthiocholine and butyrylthiocholine are in hibited in parallel by cholinesterase inhibitors, with the AChE-specif ic drug BW284c51 being the most potent; however, this drug and propidi um, a peripheral anionic site ligand, are much weaker inhibitors of th e hagfish enzyme than of Torpedo AChE. We used sequential extraction, collagenase digestion, and velocity sedimentation on sucrose gradients to determine that the AChE from the skeletal muscle of the hagfish is present in both globular and asymmetric forms. We also used the polym erase chain reaction with degenerate oligonucleotide probes and genomi c DNA to obtain a 1 kb gene fragment for hagfish AChE. The enzyme has an acyl binding site typical of other vertebrate AChE, but lacks two a romatic residues implicated in the function of the peripheral anionic subsite. We discuss the relevance of our findings to the evolution of the cholinesterases in the vertebrates.