Molecular and cellular biology of cholinesterases

Novel functions of cholinesterases in development, physiology and disease

Acetylcholinesterase (AChE; EC 3.1.1.7) is the primary terminator of nerve impulse transmission at cholinergic synapses and is believed to play an important role in neural development. Targeted deletion of four exons of the ACHE gene reduced AChE activity by half in heterozygous mutant mice and totally eliminated AChE activity in nullizygous animals. Butyrylcholinesterase (EC 3.1.1.8) activity was normal in AChE -/- mice. Although nullizygous mice were born alive and lived up to 21 days, physical development was delayed. The neuromuscular junction of 12-day-old nullizygous animals appeared normal in structure. Nullizygous mice were highly sensitive to the toxic effects of the organophosphate diisopropylfluorophosphate and to the butyrylcholinesterase-specific inhibitor bambuterol. These findings indicate that butyrylcholinesterase and possibly other enzymes are capable of compensating for some functions of AChE and that the inhibition of targets other than AChE by organophosphorus agents results in death.

Postnatal Developmental Delay and Supersensitivity to Organophosphate in Gene-Targeted Mice Lacking Acetylcholinesterase

Insect acetylcholinesterase: catalytic properties, tissue distribution and molecular forms.

We have isolated five genomic clones for human butyrylcholinesterase (BChE), using cDNA probes encoding the catalytic subunit of the hydrophilic tetramer [McTiernan et al. (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 6682-6686]. The BChE gene is at least 73 kb long and contains four exons. Exon 1 contains untranslated sequences and two potential translation initiation sites at codons -69 and -47. Exon 2 (1525 bp) contains 83% of the coding sequence for the mature protein, including the N-terminal and the active-site serine, and a third possible translation initiation site (likely functional), at codon -28. Exon 3 is 167 nucleotides long. Exon 4 (604 bp) codes for the C-terminus of the protein and the 3' untranslated region where two polyadenylation signals were identified. Intron 1 is 6.5 kb long, and the minimal sizes of introns 2 and 3 are estimated to be 32 kb each. Southern blot analysis of total human genomic DNA is in complete agreement with the gene structure established by restriction endonuclease mapping of the genomic clones: this strongly suggests that the BChE gene is present in a single copy.

Structure of the gene for human butyrylcholinesterase. Evidence for a single copy.

Cholinesterases (ChEs) hydrolyse choline esters specifically and rapidly and thus play an essential role in cholinergic transmission, e. g. at the neuromuscular junctions of vertebrates. Vertebrates possess two distinct ChEs, acetylcholinesterase (AChE, EC 3.1.1.7) and butyrylcholinesterase, also called pseudocholinesterase or non-specific cholinesterase (BuChE, EC 3.1.1.8). Both these enzymes present multiple molecular variants which possess identical catalytic activity but differ in their molecular structure and interactions. The molecular variants can be differentiated by various analytical procedures. We find it useful first to classify them as molecular forms, according to their hydrodynamic parameters (sedimentation coefficient, Stokes radius), corresponding to different quaternary structures.

J. Massoulié

Papers

Vertebrate Cholinesterases: Structure and Types of Interaction