| CHEMICAL MODIFICATION OF SULFHYDRYL GROUPS OF E.coli LEUCYL-tRNA SYNTHE-TASE AND SEQUENCING OF [~(14)C]NEM-LABELED PEPTIDES |
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| 作者姓名： | 缪枫 施建平 王应睐 |
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| 作者单位： | Shanghai Institute oj Biochemistry，Academia Sinica，Shanghai 200031，PRC，Shanghai Institute oj Biochemistry，Academia Sinica，Shanghai 200031，PRC，Shanghai Institute oj Biochemistry，Academia Sinica，Shanghai 200031，PRC |
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| 基金项目： | Supported by the National Natural Science Foundation of China and the President's Research Foundation of Academia Sinica. |
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| 摘 要： | The chemical modification of the sulfhydryl groups of E. coli Leucyl--tRNA synthetase(LeuRS) by DTNB, NEM and IAA resulted in a time-dependent loss of both amino-acid acti-vation and aminoacylation activities in parallel. The second-order reaction constants of DTNB,NEM and IAA were 1700, 150 and 0.46 mol/L~(-1) min~(-1) respectively. Chemical stoichiometryshowed that only one sulfhydryl group of LeuRS was essential for both activities. Substratesleucine and Leu-AMP protected the active sulfhydryl group from modification, suggestingthat the modified sulfhydryl group is located in or near the active site region responsiblefor amino-acid activation. ~(14)C]NEM--labeled LeuRS was subjected to tryptic digestion, andpeptides were separated and sequenced. 179 Cys~*-Asp-Thr-Leu182 was identified as the major~(14)C]NEM-labeled site in LeuRS. This result is consistent with the previous observationthat the region for Leu--AMP formation was located at the N--terminal part of LeuRS.
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CHEMICAL MODIFICATION OF SULFHYDRYL GROUPS OF E.coli LEUCYL-tRNA SYNTHE-TASE AND SEQUENCING OF [~(14)C]NEM-LABELED PEPTIDES |
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| MIAO FENG,SHI JIAN--PING,WANG YING-LAI Shanghai Institute oj Biochemistry,Academia Sinica,Shanghai ,PRC.CHEMICAL MODIFICATION OF SULFHYDRYL GROUPS OF E.coli LEUCYL-tRNA SYNTHE-TASE AND SEQUENCING OF [~(14)C]NEM-LABELED PEPTIDES[J].Science in China(Chemistry),1991(6). |
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| Authors: | MIAO FENG SHI JIAN--PING WANG YING-LAI Shanghai Institute oj Biochemistry Academia Sinica Shanghai PRC |
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| Institution: | MIAO FENG,SHI JIAN--PING,WANG YING-LAI Shanghai Institute oj Biochemistry,Academia Sinica,Shanghai 200031,PRC |
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| Abstract: | The chemical modification of the sulfhydryl groups of E. coli Leucyl--tRNA synthetase(LeuRS) by DTNB, NEM and IAA resulted in a time-dependent loss of both amino-acid acti-vation and aminoacylation activities in parallel. The second-order reaction constants of DTNB,NEM and IAA were 1700, 150 and 0.46 mol/L~(-1) min~(-1) respectively. Chemical stoichiometryshowed that only one sulfhydryl group of LeuRS was essential for both activities. Substratesleucine and Leu-AMP protected the active sulfhydryl group from modification, suggestingthat the modified sulfhydryl group is located in or near the active site region responsiblefor amino-acid activation. ~(14)C]NEM--labeled LeuRS was subjected to tryptic digestion, andpeptides were separated and sequenced. 179 Cys~*-Asp-Thr-Leu182 was identified as the major~(14)C]NEM-labeled site in LeuRS. This result is consistent with the previous observationthat the region for Leu--AMP formation was located at the N--terminal part of LeuRS. |
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| Keywords: | LeuRS chemical modification [~(14)C]NEM-labeled peptide |
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