PubMed:7000775 JSONTXT

{"target":"https://pubannotation.org/docs/sourcedb/PubMed/sourceid/7000775","sourcedb":"PubMed","sourceid":"7000775","source_url":"http://www.ncbi.nlm.nih.gov/pubmed/7000775","text":"Differential reactivity of the functional sulfhydryl groups of cysteine-32 and cysteine-35 present in the reduced form of thioredoxin from Escherichia coli.\nOnly one of the sulfhydryl groups from Cys-32 and Cys-35 in the active center of native Escherichia coli thioredoxin-(SH)2 was alkylated by excess iodoacetic acid at pH values below 8.0. Both groups reacted in the protein denatured with 4.5 M guanidine hydrochloride. The second order rate of alkylation of thioredoxin-(SH)2 with 1 eq of iodoacetic acid was pH-dependent and showed independent initial reactions of one thiolate ion with a pK value of 6.7 and a second with a pK value close to 9.0. The same pH dependence was observed for alkylation with iodoacetamide but the apparent rate constant, 107 M-1 S-1 at pH 7.2, was about 20-fold higher than the corresponding rate with iodoacetate. The sulfhydryl group with a pK value of 6.7 was shown to belong to Cys-32 by labeling thioredoxin with [14C]iodoacetic acid followed by complete alkylation with [3H]iodoacetate and amino acid sequence analysis of peptides from the active center. The abnormally low pK value of Cys-32 is suggested to arise by electrostatic influence from a positive charge on the amino group of Lys-36. A mechanism of action for thioredoxin-(SH)2 as a protein disulfide reductase has been formulated. This is based on an initial nucleophilic attack by the thiolate of Cys-32 with the formation of an unstable transient mixed disulfide involving Cys-32 and one of the sulfurs in the substrate. This is followed by a conformational change and a nucleophilic attack of Cys-35 to give the 14-membered disulfide ring in thioredoxin-S2 and the dithiol of the substrate.","tracks":[{"project":"sentences","denotations":[{"id":"T1","span":{"begin":0,"end":156},"obj":"Sentence"},{"id":"T2","span":{"begin":157,"end":343},"obj":"Sentence"},{"id":"T3","span":{"begin":344,"end":424},"obj":"Sentence"},{"id":"T4","span":{"begin":425,"end":654},"obj":"Sentence"},{"id":"T5","span":{"begin":655,"end":850},"obj":"Sentence"},{"id":"T6","span":{"begin":851,"end":1096},"obj":"Sentence"},{"id":"T7","span":{"begin":1097,"end":1236},"obj":"Sentence"},{"id":"T8","span":{"begin":1237,"end":1334},"obj":"Sentence"},{"id":"T9","span":{"begin":1335,"end":1526},"obj":"Sentence"},{"id":"T10","span":{"begin":1527,"end":1697},"obj":"Sentence"},{"id":"T1","span":{"begin":0,"end":156},"obj":"Sentence"},{"id":"T2","span":{"begin":157,"end":343},"obj":"Sentence"},{"id":"T3","span":{"begin":344,"end":424},"obj":"Sentence"},{"id":"T4","span":{"begin":425,"end":654},"obj":"Sentence"},{"id":"T5","span":{"begin":655,"end":850},"obj":"Sentence"},{"id":"T6","span":{"begin":851,"end":1096},"obj":"Sentence"},{"id":"T7","span":{"begin":1097,"end":1236},"obj":"Sentence"},{"id":"T8","span":{"begin":1237,"end":1334},"obj":"Sentence"},{"id":"T9","span":{"begin":1335,"end":1526},"obj":"Sentence"},{"id":"T10","span":{"begin":1527,"end":1697},"obj":"Sentence"}],"namespaces":[{"prefix":"_base","uri":"http://pubannotation.org/ontology/tao.owl#"}],"attributes":[{"subj":"T1","pred":"source","obj":"sentences"},{"subj":"T2","pred":"source","obj":"sentences"},{"subj":"T3","pred":"source","obj":"sentences"},{"subj":"T4","pred":"source","obj":"sentences"},{"subj":"T5","pred":"source","obj":"sentences"},{"subj":"T6","pred":"source","obj":"sentences"},{"subj":"T7","pred":"source","obj":"sentences"},{"subj":"T8","pred":"source","obj":"sentences"},{"subj":"T9","pred":"source","obj":"sentences"},{"subj":"T10","pred":"source","obj":"sentences"},{"subj":"T1","pred":"source","obj":"sentences"},{"subj":"T2","pred":"source","obj":"sentences"},{"subj":"T3","pred":"source","obj":"sentences"},{"subj":"T4","pred":"source","obj":"sentences"},{"subj":"T5","pred":"source","obj":"sentences"},{"subj":"T6","pred":"source","obj":"sentences"},{"subj":"T7","pred":"source","obj":"sentences"},{"subj":"T8","pred":"source","obj":"sentences"},{"subj":"T9","pred":"source","obj":"sentences"},{"subj":"T10","pred":"source","obj":"sentences"}]},{"project":"NCBITAXON","denotations":[{"id":"T1","span":{"begin":139,"end":155},"obj":"OrganismTaxon"},{"id":"T2","span":{"begin":245,"end":261},"obj":"OrganismTaxon"}],"attributes":[{"id":"A1","pred":"db_id","subj":"T1","obj":"562"},{"id":"A2","pred":"db_id","subj":"T2","obj":"562"},{"subj":"T1","pred":"source","obj":"NCBITAXON"},{"subj":"T2","pred":"source","obj":"NCBITAXON"}]}],"config":{"attribute types":[{"pred":"source","value type":"selection","values":[{"id":"sentences","color":"#9393ec","default":true},{"id":"NCBITAXON","color":"#adec93"}]}]}}