Functional consequences and structural interpretation of mutations of human choline acetyltransferase

Abstract

Choline acetyltransferase (ChAT; EC 2.3.1.6) catalyzes synthesis of acetylcholine from acetyl-CoA and choline in cholinergic neurons. Mutations in CHAT (MIM # 118490) cause potentially lethal congenital myasthenic syndromes associated with episodic apnea (ChAT-CMS) (MIM # 254210). Here we analyze the functional consequences of 12 missense and 1 nonsense mutations of CHAT in 11 patients. Nine of the mutations are novel. We examine expression of the recombinant missense mutants in Bosc 23 cells, determine their kinetic properties and thermal stability, and interpret the functional effects of 11 mutations in the context of the atomic structural model of human ChAT. Five mutations (p.Trp421Ser, p.Ser498Pro, p.Thr553Asn, p.Ala557Thr, p.Ser572Trp) reduce enzyme expression to <50% of wild-type. Mutations with severe kinetic effects are located in the active-site tunnel (p.Met202Arg, p.Thr553Asn and p.Ala557Thr) or adjacent to the substrate binding site (p.Ser572Trp), or exert their effect allosterically (p.Trp421Ser and p.Ile689Ser). Two mutations with milder kinetic effects (p.Val136Met, p.Ala235Thr) are also predicted to act allosterically. One mutation (p.Thr608Asn) below the nucleotide binding site of CoA enhances dissociation of AcCoA from the enzyme-substrate complex. Two mutations introducing a proline residue into an α-helix (p.Ser498Pro and p.Ser704Pro) impair the thermal stability of ChAT.

Congenital Myasthenic Syndromes in 2012

“…As indicated above under the section on “Clinical Diagnosis of the Congenital Myasthenic Syndrome”, this syndrome is associated with sudden episodes of apnea against a background of variable myasthenic symptoms [9, 10, 11•]. Most patients respond to anti-AChE therapy but few remain apneic and paralyzed since birth, and some develop cerebral atrophy after episodes of hypoxemia [11•].…”

“…Most patients respond to anti-AChE therapy but few remain apneic and paralyzed since birth, and some develop cerebral atrophy after episodes of hypoxemia [11•]. The observed mutations alter the expression or kinetic properties or structural stability of ChAT.…”

Congenital myasthenic syndromes and the neuromuscular junction

“…This could be either related to brain hypoxemia or ChAT deficiency in the brain, where it is widely expressed. Nine novel CHAT mutations were reported [29]. The variants were responsible for low expression of ChAT, alteration of the enzyme catalytic efficiency and compromise in the thermal stability of the mutant protein.…”

Inherited disorders of the neuromuscular junction: an update

“…This could be either related to brain hypoxaemia or ChAT deficiency in the brain, where it is widely expressed. Nine novel CHAT mutations were reported [37]. The variants were responsible for low expression of ChAT, alteration of the enzyme catalytic efficiency and compromise in the thermal stability of the mutant protein.…”