Abstract:
:The Escherichia coli phenylalanine-sensitive DAHP (3-deoxy-D-arabino-heptulosonate 7-phosphate) synthetase (aroG product) is one of the DAHP synthetase isozymes that catalyze the first committed step in the biosynthesis of aromatic amino acids and vitamins. Through target-directed mutagenesis of the cloned aroG on a plasmid vector, followed by screening of phenylalanine-resistant colonies, we isolated a clone (pG908) showing feedback-insensitive mutation of DAHP synthetase. The mutations were identified as a T-->A mutation at nucleotide 22, and a C-->T mutation at nucleotide 539, causing a Leu-8 to Ile-8 mutation and a Ser-180 to Phe-180 substitution, respectively. The resulting enzyme exhibited comparable enzymatic activity to the wild type, but the degree of feedback inhibition had declined from approximately 60% to less than 10% in the presence of 20 mM phenylalanine in the assay medium. Replacement of Ile-8 by Leu, and substitution of Phe-180 with Ser, Asn and Cys, using site-directed mutagenesis, demonstrated that Ser-180 is a critical residue in the feedback inhibition of AroG. The result suggests that the major role played by Ser-180 may not involve a simple charge-size effect in the Phe-binding site of the enzyme molecule, but rather may involve more complicated molecular interactions occurring in the feedback inhibition mechanisms.
journal_name
J Biochemjournal_title
Journal of biochemistryauthors
Ger YM,Chen SL,Chiang HJ,Shiuan Ddoi
10.1093/oxfordjournals.jbchem.a124657subject
Has Abstractpub_date
1994-11-01 00:00:00pages
986-90issue
5eissn
0021-924Xissn
1756-2651journal_volume
116pub_type
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journal_title:Journal of biochemistry
pub_type: 杂志文章
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更新日期:2005-12-01 00:00:00
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journal_title:Journal of biochemistry
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journal_title:Journal of biochemistry
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更新日期:2007-07-01 00:00:00
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pub_type: 杂志文章
doi:10.1093/oxfordjournals.jbchem.a131398
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journal_title:Journal of biochemistry
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journal_title:Journal of biochemistry
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journal_title:Journal of biochemistry
pub_type: 杂志文章
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pub_type: 杂志文章
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journal_title:Journal of biochemistry
pub_type: 杂志文章
doi:10.1093/oxfordjournals.jbchem.a133816
更新日期:1982-04-01 00:00:00
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pub_type: 杂志文章
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journal_title:Journal of biochemistry
pub_type: 杂志文章
doi:10.1093/oxfordjournals.jbchem.a135005
更新日期:1984-12-01 00:00:00
abstract::We can detect phospholipase A activity in non-hemolyzed sheep erythrocytes, using dilauroylglycerophosphocholine as an exogenous substrate. Only substrates such as dilauroylglycerophosphocholine, which can be incorporated into membranes, could be hydrolyzed by the enzyme, egg phosphatidylcholine being only slightly se...
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pub_type: 杂志文章
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更新日期:1983-09-01 00:00:00