{"id":{"repo_id":"unt","oai_identifier":"info:ark/67531/metadc2183"},"canonical_url":"https://search.dev.ndltd.org/etd/unt/info:ark/67531/metadc2183","repository":{"repo_id":"unt","name":"University of North Texas","base_url":"https://digital.library.unt.edu/oai/"},"display":{"title":"Nucleotide Inhibition of Glyoxalase II","abstract":"The glyoxalase system mediates the conversion of methylglyoxal, a toxic ketoaldehyde, to D-lactic acid. The system is composed of two enzymes, glyoxalase I (Glo-I) and glyoxalase II (Glo-II), and exhibits an absolute requirement for a catalytic quantity of glutathione (GSH). Glo-I catalyzes the isomerization of a hemithioacetal, formed non-enzymatically from methylglyoxal and GSH, to the corresponding a -D-hydroxyacid thioester, s-D-lactoylglutathione (SLG). Glo-II catalyzes the irreversible breakdown of SLG to D-lactate and GSH. We have observed that ATP or GTP significantly inhibits the Glo-II activity of tissue homogenates from various sources. We have developed a rapid, one step chromatography procedure to purify Glo-II such that the purified enzyme remains \"sensitive\" to inhibition by ATP or GTP (Glo-II-s). Studies indicate that inhibition of Glo-II-s by nucleotides is restricted to ATP, GTP, ADP, and GDP, with ATP appearing most effective. Kinetics studies have shown that ATP acts as a partial non-competitive inhibitor of Glo-II-s activity, and further suggest that two kinetically distinguishable forms of the enzyme exist. The sensitivity of pure Glo-II-s to nucleotide inhibition is slowly lost on storage even at -80° C. This loss is accelerated at higher temperatures or in the presence of ATP. Kinetics studies on the resultant \"insensitive\" enzyme (Glo-II-i) show that a significant reduction of the affinity of the enzyme for the substrate, SLG, occurs and further suggest that only one form of the enzyme is kinetically distinguishable after \"de-sensitization\". Tryptophan fluorescence studies of the two enzyme preparations suggest that a subtle conformational change in the enzyme has occurred during de-sensitization. We have also observed that Glo-II-i is \"resensitized\" to nucleotide inhibition after incubation in the presence of a reagent that reduces disulfide bonds. The resensitized enzyme exhibits an increased KM value similar to that of the original Glo-II-s. Kinetics studies show that ATP or GTP again act as partial non-competitive inhibitors of the resensitized enzyme and suggest that only one form of the enzyme is present. The physiological significance of the two enzyme forms is discussed.","abstract_html":"The glyoxalase system mediates the conversion of methylglyoxal, a toxic ketoaldehyde, to D-lactic acid. The system is composed of two enzymes, glyoxalase I (Glo-I) and glyoxalase II (Glo-II), and exhibits an absolute requirement for a catalytic quantity of glutathione (GSH). Glo-I catalyzes the isomerization of a hemithioacetal, formed non-enzymatically from methylglyoxal and GSH, to the corresponding a -D-hydroxyacid thioester, s-D-lactoylglutathione (SLG). Glo-II catalyzes the irreversible breakdown of SLG to D-lactate and GSH. We have observed that ATP or GTP significantly inhibits the Glo-II activity of tissue homogenates from various sources. We have developed a rapid, one step chromatography procedure to purify Glo-II such that the purified enzyme remains "sensitive" to inhibition by ATP or GTP (Glo-II-s). Studies indicate that inhibition of Glo-II-s by nucleotides is restricted to ATP, GTP, ADP, and GDP, with ATP appearing most effective. Kinetics studies have shown that ATP acts as a partial non-competitive inhibitor of Glo-II-s activity, and further suggest that two kinetically distinguishable forms of the enzyme exist. The sensitivity of pure Glo-II-s to nucleotide inhibition is slowly lost on storage even at -80° C. This loss is accelerated at higher temperatures or in the presence of ATP. Kinetics studies on the resultant "insensitive" enzyme (Glo-II-i) show that a significant reduction of the affinity of the enzyme for the substrate, SLG, occurs and further suggest that only one form of the enzyme is kinetically distinguishable after "de-sensitization". Tryptophan fluorescence studies of the two enzyme preparations suggest that a subtle conformational change in the enzyme has occurred during de-sensitization. We have also observed that Glo-II-i is "resensitized" to nucleotide inhibition after incubation in the presence of a reagent that reduces disulfide bonds. The resensitized enzyme exhibits an increased KM value similar to that of the original Glo-II-s. Kinetics studies show that ATP or GTP again act as partial non-competitive inhibitors of the resensitized enzyme and suggest that only one form of the enzyme is present. The physiological significance of the two enzyme forms is discussed.","abstract_has_math":false,"creators":["Gillis, Glen S"],"institution":"University of North Texas","degree_name":null,"degree_level":null,"degree_discipline":null,"degree_department":null,"school":null,"contributors":["Norton, Scott J.","Massarachia, Ruth Anne","Donahue, Manus J.","Chapman, Kent D."],"advisors":[],"committee_chairs":[],"committee_members":[],"year":1999,"date_issued":"1999-05","date_published":"1999-05","updated_at":"2026-07-24T05:34:52Z","subjects":["Glyoxalase.","Nucleotides.","Glutathione.","glyoxalase I","glutathione","ATP","GTP","glyoxalase II"],"languages":["English"],"rights":["Public","Copyright","Gillis, Glen S","Copyright is held by the author, unless otherwise noted. All rights reserved."],"rights_urls":[],"identifier_entries":[{"key":"dc:identifier","label":"Identifier","values":["oclc: 45003473","untcat: b2226120","https://digital.library.unt.edu/ark:/67531/metadc2183/","ark: ark:/67531/metadc2183"],"render_values":[{"text":"oclc: 45003473","href":null,"code":true},{"text":"untcat: b2226120","href":null,"code":true},{"text":"https://digital.library.unt.edu/ark:/67531/metadc2183/","href":"https://digital.library.unt.edu/ark:/67531/metadc2183/","code":true},{"text":"ark: ark:/67531/metadc2183","href":null,"code":true}]}]},"links":{"outbound_url":"https://doi.org/10.12794/metadc2183","outbound_label":"DOI","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Norton, Scott J.","Massarachia, Ruth Anne","Donahue, Manus J.","Chapman, Kent D."]},{"key":"dc:creator","label":"Author","values":["Gillis, Glen S"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["1999-05"]},{"key":"dc:publisher","label":"Institution","values":["University of North Texas"]},{"key":"dc:type","label":"Dc Type","values":["Thesis or Dissertation"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["Glyoxalase.","Nucleotides.","Glutathione.","glyoxalase I","glutathione","ATP","GTP","glyoxalase II"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["English"]},{"key":"dc:rights","label":"Dc Rights","values":["Public","Copyright","Gillis, Glen S","Copyright is held by the author, unless otherwise noted. All rights reserved."]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["oclc: 45003473","untcat: b2226120","doi: 10.12794/metadc2183","https://digital.library.unt.edu/ark:/67531/metadc2183/","ark: ark:/67531/metadc2183"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["The glyoxalase system mediates the conversion of methylglyoxal, a toxic ketoaldehyde, to D-lactic acid. The system is composed of two enzymes, glyoxalase I (Glo-I) and glyoxalase II (Glo-II), and exhibits an absolute requirement for a catalytic quantity of glutathione (GSH). Glo-I catalyzes the isomerization of a hemithioacetal, formed non-enzymatically from methylglyoxal and GSH, to the corresponding a -D-hydroxyacid thioester, s-D-lactoylglutathione (SLG). Glo-II catalyzes the irreversible breakdown of SLG to D-lactate and GSH. We have observed that ATP or GTP significantly inhibits the Glo-II activity of tissue homogenates from various sources. We have developed a rapid, one step chromatography procedure to purify Glo-II such that the purified enzyme remains \"sensitive\" to inhibition by ATP or GTP (Glo-II-s). Studies indicate that inhibition of Glo-II-s by nucleotides is restricted to ATP, GTP, ADP, and GDP, with ATP appearing most effective. Kinetics studies have shown that ATP acts as a partial non-competitive inhibitor of Glo-II-s activity, and further suggest that two kinetically distinguishable forms of the enzyme exist. The sensitivity of pure Glo-II-s to nucleotide inhibition is slowly lost on storage even at -80° C. This loss is accelerated at higher temperatures or in the presence of ATP. Kinetics studies on the resultant \"insensitive\" enzyme (Glo-II-i) show that a significant reduction of the affinity of the enzyme for the substrate, SLG, occurs and further suggest that only one form of the enzyme is kinetically distinguishable after \"de-sensitization\". Tryptophan fluorescence studies of the two enzyme preparations suggest that a subtle conformational change in the enzyme has occurred during de-sensitization. We have also observed that Glo-II-i is \"resensitized\" to nucleotide inhibition after incubation in the presence of a reagent that reduces disulfide bonds. The resensitized enzyme exhibits an increased KM value similar to that of the original Glo-II-s. Kinetics studies show that ATP or GTP again act as partial non-competitive inhibitors of the resensitized enzyme and suggest that only one form of the enzyme is present. The physiological significance of the two enzyme forms is discussed."]},{"key":"dc:format","label":"Dc Format","values":["Text"]},{"key":"dc:title","label":"Title","values":["Nucleotide Inhibition of Glyoxalase II"]}]}],"canonical_facts":{"dc:contributor":["Norton, Scott J.","Massarachia, Ruth Anne","Donahue, Manus J.","Chapman, Kent D."],"dc:creator":["Gillis, Glen S"],"dc:date":["1999-05"],"dc:description":["The glyoxalase system mediates the conversion of methylglyoxal, a toxic ketoaldehyde, to D-lactic acid. The system is composed of two enzymes, glyoxalase I (Glo-I) and glyoxalase II (Glo-II), and exhibits an absolute requirement for a catalytic quantity of glutathione (GSH). Glo-I catalyzes the isomerization of a hemithioacetal, formed non-enzymatically from methylglyoxal and GSH, to the corresponding a -D-hydroxyacid thioester, s-D-lactoylglutathione (SLG). Glo-II catalyzes the irreversible breakdown of SLG to D-lactate and GSH. We have observed that ATP or GTP significantly inhibits the Glo-II activity of tissue homogenates from various sources. We have developed a rapid, one step chromatography procedure to purify Glo-II such that the purified enzyme remains \"sensitive\" to inhibition by ATP or GTP (Glo-II-s). Studies indicate that inhibition of Glo-II-s by nucleotides is restricted to ATP, GTP, ADP, and GDP, with ATP appearing most effective. Kinetics studies have shown that ATP acts as a partial non-competitive inhibitor of Glo-II-s activity, and further suggest that two kinetically distinguishable forms of the enzyme exist. The sensitivity of pure Glo-II-s to nucleotide inhibition is slowly lost on storage even at -80° C. This loss is accelerated at higher temperatures or in the presence of ATP. Kinetics studies on the resultant \"insensitive\" enzyme (Glo-II-i) show that a significant reduction of the affinity of the enzyme for the substrate, SLG, occurs and further suggest that only one form of the enzyme is kinetically distinguishable after \"de-sensitization\". Tryptophan fluorescence studies of the two enzyme preparations suggest that a subtle conformational change in the enzyme has occurred during de-sensitization. We have also observed that Glo-II-i is \"resensitized\" to nucleotide inhibition after incubation in the presence of a reagent that reduces disulfide bonds. The resensitized enzyme exhibits an increased KM value similar to that of the original Glo-II-s. Kinetics studies show that ATP or GTP again act as partial non-competitive inhibitors of the resensitized enzyme and suggest that only one form of the enzyme is present. The physiological significance of the two enzyme forms is discussed."],"dc:format":["Text"],"dc:identifier":["oclc: 45003473","untcat: b2226120","doi: 10.12794/metadc2183","https://digital.library.unt.edu/ark:/67531/metadc2183/","ark: ark:/67531/metadc2183"],"dc:language":["English"],"dc:publisher":["University of North Texas"],"dc:rights":["Public","Copyright","Gillis, Glen S","Copyright is held by the author, unless otherwise noted. All rights reserved."],"dc:subject":["Glyoxalase.","Nucleotides.","Glutathione.","glyoxalase I","glutathione","ATP","GTP","glyoxalase II"],"dc:title":["Nucleotide Inhibition of Glyoxalase II"],"dc:type":["Thesis or Dissertation"]},"updated_at":"2026-07-24T05:34:52Z"}