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A Chatonnet

Publications and source records attributed to A Chatonnet.

30 records · Page 2Linked to original sources
PubMed1991

Use of the polymerase chain reaction for homology probing of butyrylcholinesterase from several vertebrates.

Genomic blots from man, monkey, cow, sheep, pig, rabbit, dog, rat, mouse, guinea pig, and chicken DNA were hybridized with probes derived from the four exons of the human butyrylcholinesterase gene (BCHE) (Arpagaus, M., Kott, M., Vatsis, K. P., Bartels, C. F., La Du, B. N., and Lockridge, O. (1990) Biochemistry 29, 124-131). Results showed that the BCHE gene was present in a single copy in the genome of all these vertebrates. The polymerase chain reaction was used to amplify genomic DNA from these animals with oligonucleotides derived from the human BCHE coding sequence. The amplified segment contained 423 bp of BCHE sequence including the active site serine of the enzyme (amino acid 198) and a component of the anionic site, aspartate 70. Amplification was successful for monkey, pig, cow, dog, sheep, and rabbit DNA, but unsuccessful for rat, guinea pig, mouse, and chicken DNA. Amplified segments were cloned in M13 and sequenced. The mouse sequence was obtained by sequencing a genomic clone. The highest identity of the human amino acid sequence was found with monkey (100%) and the lowest with mouse (91.5%). The sequence around the active site serine 198, Phe-Gly-Glu-Ser-Ala-Gly-Ala, was conserved in all eight animals as was the anionic site component, aspartate 70. A phylogenetic tree of mammalian butyrylcholinesterases was constructed using the partial BCHE sequences.

Amino Acid Sequence↗
PubMed1991

Structure of rabbit butyrylcholinesterase gene deduced from genomic clones and from cDNA with introns.

1. Three clones were isolated from a rabbit genomic library. They covered the entire coding sequence of the rabbit BChE gene. The positions of splice sites between exons 2, 3, and 4 are identical to those found in the human gene (Arpagaus et al., 1990). Exon 2 covers 83% of the coding sequence. This contrasts with the small size of exon 3 (167 bp) and large size of introns 2 and 3 (greater than 20 kb each). The active-site serine at position 198 is found in a highly conserved region. Aspartic acids in positions 91 and 170 are conserved in human and rabbit, and one of them could be involved in the calytic triad. Aspartic acid 70, present in the anionic site of human BChE, is also conserved in rabbit BChE. The coding sequences of human and rabbit BChE are 89% identical over 744 bp around the active-site serine. 2. In addition to the genomic clones, one cDNA clone (BNY1) was isolated. This cDNA was unusual in that it contained intronic sequences. The insert of 1 kb contained 167 coding bases homologous to the nucleotide sequence 1434 to 1600 of human cDNA and corresponded to exon 3 of the BChE gene. On each side of this coding region, consensus sequences of intron-exon boundaries were found. 3. The presence of large-size transcripts in Northern blots and the existence of a cDNA copy of unprocessed mRNA found in the BNY1 clone suggest a slow processing of transcripts. A genomic sequence unspliced in a cDNA of Torpedo AChE could give a transmembrane domain (Sikorav et al., 1988); the corresponding sequence in rabbit BChE gene, also found in a cDNA, had no homology with Torpedo AChE but could be translated in a hydrophobic C-terminal domain if maintained in mature mRNA.

Amino Acid Sequence↗
PubMed1990

Evidence for a single butyrylcholinesterase gene in individuals carrying the C5 plasma cholinesterase variant (CHE2).

DNA of 3 unrelated individuals carrying the human plasma butyrylcholinesterase C5 variant (CHE2) was isolated from white blood cells. Southern blot patterns of DNA restriction fragments probed with each of the 4 butyrylcholinesterase exons provided evidence that the production of C5 is not directed by a second butyrylcholinesterase gene. This finding supports the suggestion that the C5 variant is a hybrid enzyme resulting from the association of butyrylcholinesterase subunits with a non-cholinesterase protein.

Butyrylcholinesterase↗
PubMed1987

Brain cDNA clone for human cholinesterase.

A cDNA library from human basal ganglia was screened with oligonucleotide probes corresponding to portions of the amino acid sequence of human serum cholinesterase (EC 3.1.1.8). Five overlapping clones, representing 2.4 kilobases, were isolated. The sequenced cDNA contained 207 base pairs of coding sequence 5' to the amino terminus of the mature protein in which there were four ATG translation start sites in the same reading frame as the protein. Only the ATG coding for Met-(-28) lay within a favorable consensus sequence for functional initiators. There were 1722 base pairs of coding sequence corresponding to the protein found circulating in human serum. The amino acid sequence deduced from the cDNA exactly matched the 574 amino acid sequence of human serum cholinesterase, as previously determined by Edman degradation. Therefore, our clones represented cholinesterase (EC 3.1.1.8) rather than acetylcholinesterase (EC 3.1.1.7). It was concluded that the amino acid sequences of cholinesterase from two different tissues, human brain and human serum, were identical. Hybridization of genomic DNA blots suggested that a single gene, or very few genes, coded for cholinesterase.

Amino Acid Sequence↗
PubMed1986

Is the peptidase activity of highly purified human plasma cholinesterase due to a specific cholinesterase isoenzyme or a contaminating dipeptidylaminopeptidase?

The peptidase site of human plasma cholinesterase (butyrylcholinesterase) is distinct from its esteratic site. We found that the number of peptidase sites on an enzyme highly purified from pooled plasma is less than 0.1, as compared with 4 esteratic sites, per tetramer. However, the subunits which carry the peptidase sites are electrophoretically indistinguishable from esteratic subunits. The atypical-silent enzyme (Ea1Es1) had a much higher absolute peptidase activity when substance P was used as the substrate, and we found that the number of peptidase and esteratic sites of this enzyme was roughly the same. This suggests that the mutated esteratic site of the silent possesses a peptidase activity. The esteratic site of the usual allozyme (Eu1Eu1) has no peptidase activity towards substance P. However, a small proportion of peptidase subunits are present in all preparations of enzymes purified from the plasmas of homozygote individuals. The peptidase activity of butyrylcholinesterase might therefore correspond to a specific isoenzyme produced by an epigenetic mechanism or produced by a gene distinct from genes E1 and E2 encoding for cholinesterase subunits. However, the possibility that highly purified cholinesterase contains traces of a dipeptidylaminopeptidase cannot be completely ruled out.

Binding Sites↗
PubMed1985

Study of the peptidasic site of cholinesterase: preliminary results.

The peptidasic site of highly purified human plasma cholinesterase was investigated using active-site-directed inhibitors. Peptidase activity was assayed taking substance P as substrate. Inhibition by organophosphates indicated that the peptidasic site contained an active serine. The presence of essential histidine residues associated with serine was revealed by histidine modifications. Carboxyl group reagents showed that the active centre contained carboxyl groups in a non-polar environment. The removal of sialic acids did not alter peptidase activity. The peptidasic site of cholinesterase shared many properties with serine proteases sites and esteratic sites of cholinesterases. In addition, with the peptidasic site, as well as the esteratic site, there was always the possibility of 'aging' when inhibited by DFP or soman.

Binding Sites↗
PubMed1984

[The peptidase site of butyrylcholinesterase is distinct from the esterase site].

Highly purified human plasma butyrylcholinesterase was inhibited by reversible inhibitors of esterase activity and modified by active-site-directed irreversible inhibitors of esterases and proteases. Peptidase and esterase activities of inhibited enzyme were simultaneously essayed from rates of hydrolysis of substance P (first cleavage) and butyrylthiocholine respectively. Inhibition parameters values and rates of inactivation of the two activities provide evidence that the peptidasic site is distinct from the esteratic site.

Binding Sites↗
PubMed1983

Acetylcholinesterase molecular forms in the fast or slow muscles of the chicken and the pigeon.

Molecular forms of acetylcholinesterase (AChE) were examined in various skeletal muscles of the chicken and the pigeon. In chicken pectoralis m., AChE was found to be restricted to endplate containing segments, and no asymmetric form could be detected in aneural samples. In the chicken muscles studied, a relation has been established between globular (G1,G2,G4) forms or asymmetric (A8,A12) forms, and muscle fibre types. Asymmetric forms are preponderant in fast-twitch muscles, whereas in slow tonic muscles 80% of the AChE activity is due to globular forms. However, comparison with pigeon muscles shows that AChE chicken muscle patterns may not be generalized.

Acetylcholinesterase↗
PubMedDate not supplied

Expression of acetylcholinesterase gene during in vitro differentiation of rabbit muscle satellite cells.

We investigated the myogenic properties and the expression of acetylcholinesterase (AChE) in culture of satellite cells (SCs) isolated from slow and fast rabbit muscles. Slow SCs form myotubes more rapidly (day 9 vs day 11) than fast SCs, and differentiate further into striated and contractile fibers. AChE activity and mRNA expression are higher in SCs cultured from slow than from fast muscles, as also observed in the muscles themselves. However, the two types of SC cultures do not show obvious difference in their patterns of AChE molecular forms. Taken together, these preliminary data support the view that there might be more than one SC population in skeletal muscles.

Acetylcholinesterase↗
PubMedDate not supplied

Localization of acetylcholinesterase activity and mRNA in the rabbit embryo between 10 and 15 days.

Histoenzymatic methods and in situ hybridization were used to follow AChE expression in rabbit embryos from 10 to 15 days. Transcripts of AChE are detected at the same developmental stages in all structures where enzymatic activity is found, except in neuronal extension and the ventral part of mesonephros. AChE and BChE expression were compared. BChE transcripts are detected before BChE activity can be revealed in blood cells and mesonephros.

Acetylcholinesterase↗