Active site histidine in pig liver aminolevulic acid dehydratase modified by diethylpyrocarbonate and protected by Zn2+ Ions

1. 1. The effect of diethylpyrocarbonate (DEP) (0.1-0.35 mM) on the purified pig liver aminolevulic acid dehydratase (ALA-D) containing 0.3 g-atoms Zn/subunit, under different pHs (6.0-7.5), temperature (0-18°C) and time (0-60 min) was studied 2. 2. Three histidyl residues/subunit were modified by D...

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Autores principales:	Fukuda, H., Paredes, S.R., del C. Batlle, A.M.
Formato:	JOUR
Materias:	diethyl pyrocarbonate histidine metal porphobilinogen synthase zinc animal article binding site enzymology in vitro study liver metabolism pH swine Animal Binding Sites Diethyl Pyrocarbonate Histidine Hydrogen-Ion Concentration In Vitro Liver Metals Porphobilinogen Synthase Support, Non-U.S. Gov't Swine Zinc
Acceso en línea:	http://hdl.handle.net/20.500.12110/paper_03050491_v91_n2_p285_Fukuda
Aporte de:	Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) de Universidad de Buenos Aires

id	todo:paper_03050491_v91_n2_p285_Fukuda
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spelling	todo:paper_03050491_v91_n2_p285_Fukuda2023-10-03T15:21:21Z Active site histidine in pig liver aminolevulic acid dehydratase modified by diethylpyrocarbonate and protected by Zn2+ Ions Fukuda, H. Paredes, S.R. del C. Batlle, A.M. diethyl pyrocarbonate histidine metal porphobilinogen synthase zinc animal article binding site enzymology in vitro study liver metabolism pH swine Animal Binding Sites Diethyl Pyrocarbonate Histidine Hydrogen-Ion Concentration In Vitro Liver Metals Porphobilinogen Synthase Support, Non-U.S. Gov't Swine Zinc 1. 1. The effect of diethylpyrocarbonate (DEP) (0.1-0.35 mM) on the purified pig liver aminolevulic acid dehydratase (ALA-D) containing 0.3 g-atoms Zn/subunit, under different pHs (6.0-7.5), temperature (0-18°C) and time (0-60 min) was studied 2. 2. Three histidyl residues/subunit were modified by DEP (0.2 mM, pH 6.8), but activity was completely lost after the first one had reacted, indicating the presence of one histidine residue essential for ALA-D catalysis. Reactivation by treatment with hydroxylamine (0.7 mM, pH 7.0) confirmed that only histidine and no other nucleophile amino acids were directly involved in DEP inhibition. 3. 3. Zn ions (0.5 mM) and the substrate ALA (5-10 mM) protected against DEP inactivation, protection was dependent on pH. 4. 4. Sn, Se, Hg, Cd, Mn, Co and Pb (0.01-0.1 mM) did not significantly protect ALA-D against inactivation. 5. 5. It is concluded that the substrate and Zn binding sites and the essential histidyl residues are in close proximity in the active center. It is proposed that in the catalytic synthesis of porphobilinogen from ALA, histidine groups have the specific role of transporting protons from the aqueous media to a hydrophobic active site. © 1988. JOUR info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by/2.5/ar http://hdl.handle.net/20.500.12110/paper_03050491_v91_n2_p285_Fukuda
institution	Universidad de Buenos Aires
institution_str	I-28
repository_str	R-134
collection	Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA)
topic	diethyl pyrocarbonate histidine metal porphobilinogen synthase zinc animal article binding site enzymology in vitro study liver metabolism pH swine Animal Binding Sites Diethyl Pyrocarbonate Histidine Hydrogen-Ion Concentration In Vitro Liver Metals Porphobilinogen Synthase Support, Non-U.S. Gov't Swine Zinc
spellingShingle	diethyl pyrocarbonate histidine metal porphobilinogen synthase zinc animal article binding site enzymology in vitro study liver metabolism pH swine Animal Binding Sites Diethyl Pyrocarbonate Histidine Hydrogen-Ion Concentration In Vitro Liver Metals Porphobilinogen Synthase Support, Non-U.S. Gov't Swine Zinc Fukuda, H. Paredes, S.R. del C. Batlle, A.M. Active site histidine in pig liver aminolevulic acid dehydratase modified by diethylpyrocarbonate and protected by Zn2+ Ions
topic_facet	diethyl pyrocarbonate histidine metal porphobilinogen synthase zinc animal article binding site enzymology in vitro study liver metabolism pH swine Animal Binding Sites Diethyl Pyrocarbonate Histidine Hydrogen-Ion Concentration In Vitro Liver Metals Porphobilinogen Synthase Support, Non-U.S. Gov't Swine Zinc
description	1. 1. The effect of diethylpyrocarbonate (DEP) (0.1-0.35 mM) on the purified pig liver aminolevulic acid dehydratase (ALA-D) containing 0.3 g-atoms Zn/subunit, under different pHs (6.0-7.5), temperature (0-18°C) and time (0-60 min) was studied 2. 2. Three histidyl residues/subunit were modified by DEP (0.2 mM, pH 6.8), but activity was completely lost after the first one had reacted, indicating the presence of one histidine residue essential for ALA-D catalysis. Reactivation by treatment with hydroxylamine (0.7 mM, pH 7.0) confirmed that only histidine and no other nucleophile amino acids were directly involved in DEP inhibition. 3. 3. Zn ions (0.5 mM) and the substrate ALA (5-10 mM) protected against DEP inactivation, protection was dependent on pH. 4. 4. Sn, Se, Hg, Cd, Mn, Co and Pb (0.01-0.1 mM) did not significantly protect ALA-D against inactivation. 5. 5. It is concluded that the substrate and Zn binding sites and the essential histidyl residues are in close proximity in the active center. It is proposed that in the catalytic synthesis of porphobilinogen from ALA, histidine groups have the specific role of transporting protons from the aqueous media to a hydrophobic active site. © 1988.
format	JOUR
author	Fukuda, H. Paredes, S.R. del C. Batlle, A.M.
author_facet	Fukuda, H. Paredes, S.R. del C. Batlle, A.M.
author_sort	Fukuda, H.
title	Active site histidine in pig liver aminolevulic acid dehydratase modified by diethylpyrocarbonate and protected by Zn2+ Ions
title_short	Active site histidine in pig liver aminolevulic acid dehydratase modified by diethylpyrocarbonate and protected by Zn2+ Ions
title_full	Active site histidine in pig liver aminolevulic acid dehydratase modified by diethylpyrocarbonate and protected by Zn2+ Ions
title_fullStr	Active site histidine in pig liver aminolevulic acid dehydratase modified by diethylpyrocarbonate and protected by Zn2+ Ions
title_full_unstemmed	Active site histidine in pig liver aminolevulic acid dehydratase modified by diethylpyrocarbonate and protected by Zn2+ Ions
title_sort	active site histidine in pig liver aminolevulic acid dehydratase modified by diethylpyrocarbonate and protected by zn2+ ions
url	http://hdl.handle.net/20.500.12110/paper_03050491_v91_n2_p285_Fukuda
work_keys_str_mv	AT fukudah activesitehistidineinpigliveraminolevulicaciddehydratasemodifiedbydiethylpyrocarbonateandprotectedbyzn2ions AT paredessr activesitehistidineinpigliveraminolevulicaciddehydratasemodifiedbydiethylpyrocarbonateandprotectedbyzn2ions AT delcbatlleam activesitehistidineinpigliveraminolevulicaciddehydratasemodifiedbydiethylpyrocarbonateandprotectedbyzn2ions
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Active site histidine in pig liver aminolevulic acid dehydratase modified by diethylpyrocarbonate and protected by Zn2+ Ions

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