Acetaminophen and phenol: Substrates for both a thermostable and a thermolabile form of human platelet phenol sulfotransferase

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Abstract

Human platelet homogenates were used to test the hypotheses that multiple forms of platelet phenol sulfotransferase (PST) with differing physical properties and substrate specificities might exist and that some compounds might serve as substrates for more than one form of platelet PST activity. PST thermal stability was determined in platelet homogenates preheated for 15 min at temperatures varying from 37-49°. When dopamine was used as a substrate, 50% of the enzyme activity was inactivated at approximately 39.5°C. However, PST activity measured with 100 μM phenol as substrate was 50% inactivated at approximately 44°C. Acetaminophen (16 mM) behaved as if it might serve as a substrate for both the thermolabile and the thermostable PST activities. Kinetic experiments were performed with platelet homogenates and phenol showed a biphasic double inverse plot with two apparent K(m) values of 16 and 638 μM. Thermal treatment of platelet homogenates that caused inactivation of the thermolabile PST activity resulted in loss of the portion of the double inverse plot that gave the higher apparent K(m) value. These results suggested that, depending on the concentration tested, phenol might serve predominantly as a substrate for either the thermolabile or the thermostable form of human platelet PST. When the hypothesis that phenol might serve as a substrate for either the thermolabile or thermostable form of human platelet PST was tested further, it was found that the enzyme activity measured with 62.5 μM phenol was thermostable whereas that measured with 4 mM phenol was thermolabile. PST activity measured with 500 μM phenol was intermediate in thermal stability. Thermal inactivation studies performed by heating platelet homogenates at 43°C for variable time periods indicated that acetaminophen was also able to serve as a substrate for both the thermolabile and thermostable forms of human platelet PST activity. The results of experiments in which PST activity was measured in platelet homogenates from 23 individual subjects were also compatible with the conclusion that phenol and acetaminophen were able to serve as substrates for both the thermostable and thermolabile forms of human platelet PST activity and that the activities of these two forms of the enzyme were regulated independently.

Original languageEnglish (US)
Pages (from-to)43-51
Number of pages9
JournalJournal of Pharmacology and Experimental Therapeutics
Volume221
Issue number1
StatePublished - 1982

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Arylsulfotransferase
Acetaminophen
Phenol
Blood Platelets
Hot Temperature
Enzymes
Substrate Specificity
Heating

ASJC Scopus subject areas

  • Pharmacology

Cite this

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title = "Acetaminophen and phenol: Substrates for both a thermostable and a thermolabile form of human platelet phenol sulfotransferase",
abstract = "Human platelet homogenates were used to test the hypotheses that multiple forms of platelet phenol sulfotransferase (PST) with differing physical properties and substrate specificities might exist and that some compounds might serve as substrates for more than one form of platelet PST activity. PST thermal stability was determined in platelet homogenates preheated for 15 min at temperatures varying from 37-49°. When dopamine was used as a substrate, 50{\%} of the enzyme activity was inactivated at approximately 39.5°C. However, PST activity measured with 100 μM phenol as substrate was 50{\%} inactivated at approximately 44°C. Acetaminophen (16 mM) behaved as if it might serve as a substrate for both the thermolabile and the thermostable PST activities. Kinetic experiments were performed with platelet homogenates and phenol showed a biphasic double inverse plot with two apparent K(m) values of 16 and 638 μM. Thermal treatment of platelet homogenates that caused inactivation of the thermolabile PST activity resulted in loss of the portion of the double inverse plot that gave the higher apparent K(m) value. These results suggested that, depending on the concentration tested, phenol might serve predominantly as a substrate for either the thermolabile or the thermostable form of human platelet PST. When the hypothesis that phenol might serve as a substrate for either the thermolabile or thermostable form of human platelet PST was tested further, it was found that the enzyme activity measured with 62.5 μM phenol was thermostable whereas that measured with 4 mM phenol was thermolabile. PST activity measured with 500 μM phenol was intermediate in thermal stability. Thermal inactivation studies performed by heating platelet homogenates at 43°C for variable time periods indicated that acetaminophen was also able to serve as a substrate for both the thermolabile and thermostable forms of human platelet PST activity. The results of experiments in which PST activity was measured in platelet homogenates from 23 individual subjects were also compatible with the conclusion that phenol and acetaminophen were able to serve as substrates for both the thermostable and thermolabile forms of human platelet PST activity and that the activities of these two forms of the enzyme were regulated independently.",
author = "C. Reiter and Weinshilboum, {Richard M}",
year = "1982",
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T2 - Substrates for both a thermostable and a thermolabile form of human platelet phenol sulfotransferase

AU - Reiter, C.

AU - Weinshilboum, Richard M

PY - 1982

Y1 - 1982

N2 - Human platelet homogenates were used to test the hypotheses that multiple forms of platelet phenol sulfotransferase (PST) with differing physical properties and substrate specificities might exist and that some compounds might serve as substrates for more than one form of platelet PST activity. PST thermal stability was determined in platelet homogenates preheated for 15 min at temperatures varying from 37-49°. When dopamine was used as a substrate, 50% of the enzyme activity was inactivated at approximately 39.5°C. However, PST activity measured with 100 μM phenol as substrate was 50% inactivated at approximately 44°C. Acetaminophen (16 mM) behaved as if it might serve as a substrate for both the thermolabile and the thermostable PST activities. Kinetic experiments were performed with platelet homogenates and phenol showed a biphasic double inverse plot with two apparent K(m) values of 16 and 638 μM. Thermal treatment of platelet homogenates that caused inactivation of the thermolabile PST activity resulted in loss of the portion of the double inverse plot that gave the higher apparent K(m) value. These results suggested that, depending on the concentration tested, phenol might serve predominantly as a substrate for either the thermolabile or the thermostable form of human platelet PST. When the hypothesis that phenol might serve as a substrate for either the thermolabile or thermostable form of human platelet PST was tested further, it was found that the enzyme activity measured with 62.5 μM phenol was thermostable whereas that measured with 4 mM phenol was thermolabile. PST activity measured with 500 μM phenol was intermediate in thermal stability. Thermal inactivation studies performed by heating platelet homogenates at 43°C for variable time periods indicated that acetaminophen was also able to serve as a substrate for both the thermolabile and thermostable forms of human platelet PST activity. The results of experiments in which PST activity was measured in platelet homogenates from 23 individual subjects were also compatible with the conclusion that phenol and acetaminophen were able to serve as substrates for both the thermostable and thermolabile forms of human platelet PST activity and that the activities of these two forms of the enzyme were regulated independently.

AB - Human platelet homogenates were used to test the hypotheses that multiple forms of platelet phenol sulfotransferase (PST) with differing physical properties and substrate specificities might exist and that some compounds might serve as substrates for more than one form of platelet PST activity. PST thermal stability was determined in platelet homogenates preheated for 15 min at temperatures varying from 37-49°. When dopamine was used as a substrate, 50% of the enzyme activity was inactivated at approximately 39.5°C. However, PST activity measured with 100 μM phenol as substrate was 50% inactivated at approximately 44°C. Acetaminophen (16 mM) behaved as if it might serve as a substrate for both the thermolabile and the thermostable PST activities. Kinetic experiments were performed with platelet homogenates and phenol showed a biphasic double inverse plot with two apparent K(m) values of 16 and 638 μM. Thermal treatment of platelet homogenates that caused inactivation of the thermolabile PST activity resulted in loss of the portion of the double inverse plot that gave the higher apparent K(m) value. These results suggested that, depending on the concentration tested, phenol might serve predominantly as a substrate for either the thermolabile or the thermostable form of human platelet PST. When the hypothesis that phenol might serve as a substrate for either the thermolabile or thermostable form of human platelet PST was tested further, it was found that the enzyme activity measured with 62.5 μM phenol was thermostable whereas that measured with 4 mM phenol was thermolabile. PST activity measured with 500 μM phenol was intermediate in thermal stability. Thermal inactivation studies performed by heating platelet homogenates at 43°C for variable time periods indicated that acetaminophen was also able to serve as a substrate for both the thermolabile and thermostable forms of human platelet PST activity. The results of experiments in which PST activity was measured in platelet homogenates from 23 individual subjects were also compatible with the conclusion that phenol and acetaminophen were able to serve as substrates for both the thermostable and thermolabile forms of human platelet PST activity and that the activities of these two forms of the enzyme were regulated independently.

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