A Simple Micromethod for Measurement of CoASH and Its Use in Measuring Intracellular Levels of CoASH and Short Chain Acyl-CoAs in Eschevichia coli K12 Cells(Microbiology & Fermentation Industry)
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概要
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A simple micromethod for measurement of CoASH was established. CoASH was quantitatively converted to acetyl-CoA with phosphate acetyltransferase (EC 2.3.1.8), then acetyl-CoA originated from CoASH was measuring by the acyl-CoA cycling method using malonate CoA-transferase (EC 2.8.3.3). This method is highly sensitive and can detect 10^<-12>mol of CoASH without any loss of CoA-thioesters, and it is specific for CoASH and 3'-dephospho-CoA. The established method was useful to define the rapid changes in the size and composition of in vivo CoA pool (CoASH, acetyl-CoA, and malonyl-CoA) in Escherichia call K12. It was found that the CoA pool changed rapidly and dramatically in a minute order in ranges of CoASH; 40-300μM (0.11-0.80 nmol/mg dry wt.), acetyl-CoA; 40-350μ_M (0.12-0.93 nmol/mg dry wt.), and malonyl-CoA; 0-290μM (0-0.79 nmol/mg dry wt.). When glucose in the medium was depleted, CoASH increased to be the major component (82%) of the CoA pool, while when sufficient amount of glucose (28m_M) was provided, CoASH was reduced to a minor component (15%). In contrast with this, acetyl-CoA increased to the maximal level of 350μ_M (0.93 nmol/mg dry wt.) and to be the major component (66%) 10min after 28m_M glucose was added. A remarkable demonstration was achieved in confirmation of the reverse relationship between CoASH and short chain acyl-CoAs, particularly acetyl-CoA, of which the in vivo behavior has been mere conjecture until now.
- 社団法人日本農芸化学会の論文
- 1991-01-23
著者
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Chohnan Shigeru
Department Of Agricultural Chemistry Faculty Of Agriculture Ibaraki University
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TAKAMURA YOSHICHIKA
Department of Bioresource Science, Faculty of Agriculture, Ibaraki University
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Takamura Yoshichika
Department Of Agricultural Chemistry Faculty Of Agriculture Ibaraki University
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