Comparison of atypical and usual human serum cholinesterase. Purification, number of active sites, substrate affinity, and turnover number

Oksana Lockridge, B. N. La Du

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Abstract

Atypical and usual human serum cholinesterases were purified and studied with the fluorescent probe, N-methyl(7-dimethylcarbamoxy) quinolinium iodide. Four active sites per tetramer were found in each enzyme. The turnover numbers of usual and atypical cholinesterases were the same: 15,000 μmol of benzoylcholine hydrolyzed/min/μmol of active site: 48,000 min-1 for o-nitrophenylbutyrate; and 0.0025 min-1 for N-methyl-(7-dimethylcarbamoxy) quinolinium iodide. They had identical rate constants for carbamylation, (5.0 min-1) and for decarbamylation (0.15 h-1). The major difference between the two genetically determined forms of the enzyme was substrate affinity, K(d) being 0.16 mM for usual and 5.4 mM for atypical cholinesterase, for the fluorescent probe substrate. K(m) for the uncharged ester, o-nitrophenylbutyrate, was 0.14 mM for both enzymes, whereas K(m) for benzoylcholine was 0.005 mM for usual and 0.024 mM for atypical cholinesterase. We interpret these data to mean that the two enzymes differ only in the structure of their anionic site.

Original languageEnglish (US)
Pages (from-to)361-366
Number of pages6
JournalJournal of Biological Chemistry
Volume253
Issue number2
StatePublished - Jan 1 1978

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Cholinesterases
Purification
Benzoylcholine
Catalytic Domain
Iodides
Substrates
Enzymes
Serum
Fluorescent Dyes
Rate constants
Esters
7-(dimethylcarbamoyloxy)-N-methylquinolinium

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

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title = "Comparison of atypical and usual human serum cholinesterase. Purification, number of active sites, substrate affinity, and turnover number",
abstract = "Atypical and usual human serum cholinesterases were purified and studied with the fluorescent probe, N-methyl(7-dimethylcarbamoxy) quinolinium iodide. Four active sites per tetramer were found in each enzyme. The turnover numbers of usual and atypical cholinesterases were the same: 15,000 μmol of benzoylcholine hydrolyzed/min/μmol of active site: 48,000 min-1 for o-nitrophenylbutyrate; and 0.0025 min-1 for N-methyl-(7-dimethylcarbamoxy) quinolinium iodide. They had identical rate constants for carbamylation, (5.0 min-1) and for decarbamylation (0.15 h-1). The major difference between the two genetically determined forms of the enzyme was substrate affinity, K(d) being 0.16 mM for usual and 5.4 mM for atypical cholinesterase, for the fluorescent probe substrate. K(m) for the uncharged ester, o-nitrophenylbutyrate, was 0.14 mM for both enzymes, whereas K(m) for benzoylcholine was 0.005 mM for usual and 0.024 mM for atypical cholinesterase. We interpret these data to mean that the two enzymes differ only in the structure of their anionic site.",
author = "Oksana Lockridge and {La Du}, {B. N.}",
year = "1978",
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AU - Lockridge, Oksana

AU - La Du, B. N.

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Y1 - 1978/1/1

N2 - Atypical and usual human serum cholinesterases were purified and studied with the fluorescent probe, N-methyl(7-dimethylcarbamoxy) quinolinium iodide. Four active sites per tetramer were found in each enzyme. The turnover numbers of usual and atypical cholinesterases were the same: 15,000 μmol of benzoylcholine hydrolyzed/min/μmol of active site: 48,000 min-1 for o-nitrophenylbutyrate; and 0.0025 min-1 for N-methyl-(7-dimethylcarbamoxy) quinolinium iodide. They had identical rate constants for carbamylation, (5.0 min-1) and for decarbamylation (0.15 h-1). The major difference between the two genetically determined forms of the enzyme was substrate affinity, K(d) being 0.16 mM for usual and 5.4 mM for atypical cholinesterase, for the fluorescent probe substrate. K(m) for the uncharged ester, o-nitrophenylbutyrate, was 0.14 mM for both enzymes, whereas K(m) for benzoylcholine was 0.005 mM for usual and 0.024 mM for atypical cholinesterase. We interpret these data to mean that the two enzymes differ only in the structure of their anionic site.

AB - Atypical and usual human serum cholinesterases were purified and studied with the fluorescent probe, N-methyl(7-dimethylcarbamoxy) quinolinium iodide. Four active sites per tetramer were found in each enzyme. The turnover numbers of usual and atypical cholinesterases were the same: 15,000 μmol of benzoylcholine hydrolyzed/min/μmol of active site: 48,000 min-1 for o-nitrophenylbutyrate; and 0.0025 min-1 for N-methyl-(7-dimethylcarbamoxy) quinolinium iodide. They had identical rate constants for carbamylation, (5.0 min-1) and for decarbamylation (0.15 h-1). The major difference between the two genetically determined forms of the enzyme was substrate affinity, K(d) being 0.16 mM for usual and 5.4 mM for atypical cholinesterase, for the fluorescent probe substrate. K(m) for the uncharged ester, o-nitrophenylbutyrate, was 0.14 mM for both enzymes, whereas K(m) for benzoylcholine was 0.005 mM for usual and 0.024 mM for atypical cholinesterase. We interpret these data to mean that the two enzymes differ only in the structure of their anionic site.

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