期刊
ARCHIVES OF PHARMACAL RESEARCH
卷 28, 期 5, 页码 561-565出版社
PHARMACEUTICAL SOC KOREA
DOI: 10.1007/BF02977759
关键词
arylsulfate sulfotransferase; Enterobacter amnigenus; sodium dodecyl sulfate-polyacrylamide gel electrophoresis; disulfide bond; site-directed mutagenesis; DsbA; DsbB
Bacterial arylsulfate sulfotransferase (ASST) catalyzes the transfer of sulfate group from a phenyl sulfate ester to a phenolic acceptor. The promoter region and the transcription start sites of Enterobacter amnigenus astA have been determined by primer extension analysis. Northern blot analysis resolved two mRNA species with lengths of 3.3 and 2.0 kb, which correspond to the distances between the transcriptional initiation sites and the two inverted repeat sequences (IRSs). By length, the 3.3 kb RNA could comprise the three-gene (astA with dsbA and dsbB) operon. ASST has three highly conserved cysteine residues. Reducing and non-reducing SIDS-PAGE and activity staining showed that disulfide bond is needed for the activity of the enzyme. To identify the cysteine residues responsible for the disulfide bond formation, a series of Cys to Ser mutants has been constructed and the enzymatic activity was measured. Based on the results, we assumed that the first cysteine (Cys349) might be involved in disulfide bond mainly with the second cysteine (Cys445) and result in active conformation.
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