Qian Wang, Aleksey Aleshintsev, Kamal Rai, Eric Jin and Rupal Gupta*,
{"title":"通过具有抑制 pKa 的精氨酸进行的质子转移介导了龙胆二酸酯和水杨酸酯二氧合酶的催化作用","authors":"Qian Wang, Aleksey Aleshintsev, Kamal Rai, Eric Jin and Rupal Gupta*, ","doi":"10.1021/acs.jpcb.4c03164","DOIUrl":null,"url":null,"abstract":"<p >Gentisate and salicylate 1,2-dioxygenases (GDO and SDO) facilitate aerobic degradation of aromatic rings by inserting both atoms of dioxygen into their substrates, thereby participating in global carbon cycling. The role of acid–base catalysts in the reaction cycles of these enzymes is debatable. We present evidence of the participation of a proton shuffler during catalysis by GDO and SDO. The pH dependence of Michaelis–Menten parameters demonstrates that a single proton transfer is mandatory for the catalysis. Measurements at variable temperatures and pHs were used to determine the standard enthalpy of ionization (Δ<i>H</i><sub>ion</sub>°) of 51 kJ/mol for the proton transfer event. Although the observed apparent p<i>K</i><sub>a</sub> in the range of 6.0–7.0 for substrates of both enzymes is highly suggestive of a histidine residue, Δ<i>H</i><sub>ion</sub>° establishes an arginine residue as the likely proton source, providing phylogenetic relevance for this strictly conserved residue in the GDO family. We propose that the atypical 3-histidine ferrous binding scaffold of GDOs contributes to the suppression of arginine p<i>K</i><sub>a</sub> and provides support for this argument by employing a 2-histidine-1-carboxylate variant of the enzyme that exhibits elevated p<i>K</i><sub>a</sub>. A reaction mechanism considering the role of the proton source in stabilizing key reaction intermediates is proposed.</p>","PeriodicalId":60,"journal":{"name":"The Journal of Physical Chemistry B","volume":null,"pages":null},"PeriodicalIF":2.8000,"publicationDate":"2024-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://pubs.acs.org/doi/epdf/10.1021/acs.jpcb.4c03164","citationCount":"0","resultStr":"{\"title\":\"Proton Transfer via Arginine with Suppressed pKa Mediates Catalysis by Gentisate and Salicylate Dioxygenase\",\"authors\":\"Qian Wang, Aleksey Aleshintsev, Kamal Rai, Eric Jin and Rupal Gupta*, \",\"doi\":\"10.1021/acs.jpcb.4c03164\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p >Gentisate and salicylate 1,2-dioxygenases (GDO and SDO) facilitate aerobic degradation of aromatic rings by inserting both atoms of dioxygen into their substrates, thereby participating in global carbon cycling. The role of acid–base catalysts in the reaction cycles of these enzymes is debatable. We present evidence of the participation of a proton shuffler during catalysis by GDO and SDO. The pH dependence of Michaelis–Menten parameters demonstrates that a single proton transfer is mandatory for the catalysis. Measurements at variable temperatures and pHs were used to determine the standard enthalpy of ionization (Δ<i>H</i><sub>ion</sub>°) of 51 kJ/mol for the proton transfer event. Although the observed apparent p<i>K</i><sub>a</sub> in the range of 6.0–7.0 for substrates of both enzymes is highly suggestive of a histidine residue, Δ<i>H</i><sub>ion</sub>° establishes an arginine residue as the likely proton source, providing phylogenetic relevance for this strictly conserved residue in the GDO family. We propose that the atypical 3-histidine ferrous binding scaffold of GDOs contributes to the suppression of arginine p<i>K</i><sub>a</sub> and provides support for this argument by employing a 2-histidine-1-carboxylate variant of the enzyme that exhibits elevated p<i>K</i><sub>a</sub>. A reaction mechanism considering the role of the proton source in stabilizing key reaction intermediates is proposed.</p>\",\"PeriodicalId\":60,\"journal\":{\"name\":\"The Journal of Physical Chemistry B\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":2.8000,\"publicationDate\":\"2024-07-09\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://pubs.acs.org/doi/epdf/10.1021/acs.jpcb.4c03164\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"The Journal of Physical Chemistry B\",\"FirstCategoryId\":\"1\",\"ListUrlMain\":\"https://pubs.acs.org/doi/10.1021/acs.jpcb.4c03164\",\"RegionNum\":2,\"RegionCategory\":\"化学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q3\",\"JCRName\":\"CHEMISTRY, PHYSICAL\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"The Journal of Physical Chemistry B","FirstCategoryId":"1","ListUrlMain":"https://pubs.acs.org/doi/10.1021/acs.jpcb.4c03164","RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"CHEMISTRY, PHYSICAL","Score":null,"Total":0}
Proton Transfer via Arginine with Suppressed pKa Mediates Catalysis by Gentisate and Salicylate Dioxygenase
Gentisate and salicylate 1,2-dioxygenases (GDO and SDO) facilitate aerobic degradation of aromatic rings by inserting both atoms of dioxygen into their substrates, thereby participating in global carbon cycling. The role of acid–base catalysts in the reaction cycles of these enzymes is debatable. We present evidence of the participation of a proton shuffler during catalysis by GDO and SDO. The pH dependence of Michaelis–Menten parameters demonstrates that a single proton transfer is mandatory for the catalysis. Measurements at variable temperatures and pHs were used to determine the standard enthalpy of ionization (ΔHion°) of 51 kJ/mol for the proton transfer event. Although the observed apparent pKa in the range of 6.0–7.0 for substrates of both enzymes is highly suggestive of a histidine residue, ΔHion° establishes an arginine residue as the likely proton source, providing phylogenetic relevance for this strictly conserved residue in the GDO family. We propose that the atypical 3-histidine ferrous binding scaffold of GDOs contributes to the suppression of arginine pKa and provides support for this argument by employing a 2-histidine-1-carboxylate variant of the enzyme that exhibits elevated pKa. A reaction mechanism considering the role of the proton source in stabilizing key reaction intermediates is proposed.
期刊介绍:
An essential criterion for acceptance of research articles in the journal is that they provide new physical insight. Please refer to the New Physical Insights virtual issue on what constitutes new physical insight. Manuscripts that are essentially reporting data or applications of data are, in general, not suitable for publication in JPC B.