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Article: Further studies on the alkylation of the histidine residues at the active site of pancreatic ribonuclease.

TitleFurther studies on the alkylation of the histidine residues at the active site of pancreatic ribonuclease.
Authors
Issue Date1968
PublisherAmerican Society for Biochemistry and Molecular Biology, Inc. The Journal's web site is located at http://www.jbc.org/
Citation
Journal Of Biological Chemistry, 1968, v. 243 n. 23, p. 6167-6170 How to Cite?
AbstractThe chemistry of the active site of bovine pancreatic ribonuclease has been further examined by study of the reaction of iodoacetic acid with histidine residues 12 and 119 in derivatives of the protein. In the native enzyme, the stereospecific alkylation of the two imidazole rings is a mutually exclusive, electrostatically oriented reaction leading to a 7:1 ratio of substitution at residues 119 and 12. When inactive des-(121-124)-ribonuclease, prepared by limited peptic hydrolysis according to the method of Anfinsen, is exposed to iodoacetate at pH 5.5, alkylation at histidine-119 is nearly abolished but the reaction at histidine-12 is accelerated. If, as has been assumed for the native enzyme, 1 protonated histidine residue orients the reagent for the alkylation of the unprotonated form of the other, this result with the derivative is interpretable in terms of an increase in the pK of the imidazole ring of histidine-119 accompanying the removal of -Asp-Ala-Ser-Val from the carboxyl end of the chain. In turn, this change in pK would alter the precise interaction between the two imidazole rings which are essential for catalytic action; such a shift could account for the inactivity of the des-(121-124)-derivative. To support this interpretation of the alkylation of des-(121-124)-ribonuclease, further information on the alkylation reaction itself was sought. The influence of neighboring groups on the course of the alkylation of ribonuclease has been studied with two derivatives in which lysine residues have been modified. When the positive charge on lysine-7 is neutralized by acetylation of the S-peptide, by the procedure of Richards and Vithayathil, the resulting active Nα, ε-diAcLys-1-,Nε-Lys(Ac)-7-ribonuclease S is alkylated by iodoacetate mainly on histidine-119, as in the unaltered enzyme. The alkylation also proceeds similarly with the inactive ε-Lys(DNP)-41-ribonuclease. These results eliminate the possibility that the positive charges on the ε-NH2 groups of lysine-7 and lysine-41 have a role in the orientation of ICH2COO-.
Persistent Identifierhttp://hdl.handle.net/10722/167417
ISSN
2015 Impact Factor: 4.258
2015 SCImago Journal Rankings: 3.151

 

DC FieldValueLanguage
dc.contributor.authorLin, MCen_US
dc.contributor.authorStein, WHen_US
dc.contributor.authorMoore, Sen_US
dc.date.accessioned2012-10-08T03:06:44Z-
dc.date.available2012-10-08T03:06:44Z-
dc.date.issued1968en_US
dc.identifier.citationJournal Of Biological Chemistry, 1968, v. 243 n. 23, p. 6167-6170en_US
dc.identifier.issn0021-9258en_US
dc.identifier.urihttp://hdl.handle.net/10722/167417-
dc.description.abstractThe chemistry of the active site of bovine pancreatic ribonuclease has been further examined by study of the reaction of iodoacetic acid with histidine residues 12 and 119 in derivatives of the protein. In the native enzyme, the stereospecific alkylation of the two imidazole rings is a mutually exclusive, electrostatically oriented reaction leading to a 7:1 ratio of substitution at residues 119 and 12. When inactive des-(121-124)-ribonuclease, prepared by limited peptic hydrolysis according to the method of Anfinsen, is exposed to iodoacetate at pH 5.5, alkylation at histidine-119 is nearly abolished but the reaction at histidine-12 is accelerated. If, as has been assumed for the native enzyme, 1 protonated histidine residue orients the reagent for the alkylation of the unprotonated form of the other, this result with the derivative is interpretable in terms of an increase in the pK of the imidazole ring of histidine-119 accompanying the removal of -Asp-Ala-Ser-Val from the carboxyl end of the chain. In turn, this change in pK would alter the precise interaction between the two imidazole rings which are essential for catalytic action; such a shift could account for the inactivity of the des-(121-124)-derivative. To support this interpretation of the alkylation of des-(121-124)-ribonuclease, further information on the alkylation reaction itself was sought. The influence of neighboring groups on the course of the alkylation of ribonuclease has been studied with two derivatives in which lysine residues have been modified. When the positive charge on lysine-7 is neutralized by acetylation of the S-peptide, by the procedure of Richards and Vithayathil, the resulting active Nα, ε-diAcLys-1-,Nε-Lys(Ac)-7-ribonuclease S is alkylated by iodoacetate mainly on histidine-119, as in the unaltered enzyme. The alkylation also proceeds similarly with the inactive ε-Lys(DNP)-41-ribonuclease. These results eliminate the possibility that the positive charges on the ε-NH2 groups of lysine-7 and lysine-41 have a role in the orientation of ICH2COO-.-
dc.languageengen_US
dc.publisherAmerican Society for Biochemistry and Molecular Biology, Inc. The Journal's web site is located at http://www.jbc.org/en_US
dc.relation.ispartofJournal of Biological Chemistryen_US
dc.subject.meshAlkylationen_US
dc.subject.meshAmino Acids - Analysisen_US
dc.subject.meshAnimalsen_US
dc.subject.meshBinding Sitesen_US
dc.subject.meshCattleen_US
dc.subject.meshHistidineen_US
dc.subject.meshHydrogen-Ion Concentrationen_US
dc.subject.meshIodoacetatesen_US
dc.subject.meshLysineen_US
dc.subject.meshPancreas - Enzymologyen_US
dc.subject.meshRibonucleasesen_US
dc.titleFurther studies on the alkylation of the histidine residues at the active site of pancreatic ribonuclease.en_US
dc.typeArticleen_US
dc.identifier.emailLin, MC:mcllin@hkucc.hku.hken_US
dc.identifier.authorityLin, MC=rp00746en_US
dc.description.naturelink_to_OA_fulltexten_US
dc.identifier.pmid5723460-
dc.identifier.scopuseid_2-s2.0-0014410341en_US
dc.identifier.volume243en_US
dc.identifier.issue23en_US
dc.identifier.spage6167en_US
dc.identifier.epage6170en_US
dc.publisher.placeUnited Statesen_US
dc.identifier.scopusauthoridLin, MC=7404816359en_US
dc.identifier.scopusauthoridStein, WH=7201784892en_US
dc.identifier.scopusauthoridMoore, S=7403537387en_US

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