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KMID : 0364819880260030197
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Korean Journal of Microbiology
1988 Volume.26 No. 3 p.197 ~ p.206
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Characterization of the Outer Membrance-Associated 2-Furaldehyde Dehydrogenase from Klebsiella pneumoniae
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Lee, June-Woo/이준우
Kang, Sa-Ouk/Hah, Yung-Chil/Han, Hong-Eui/강사욱/하영칠/한홍의
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Abstract
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1
An outer membrane-associated 2-furalehyde dehydrogenase, catalyzing the oxidation of 2-furaldebyde to 2-furoic add from Kkbsieia pneumoniae was purified to homogeneity and characterized. The enzyme showed its highly specific dependency on ,9-NAD+. Enzyme activity was monitored during purification by using substrate 2-furaldehyde and coenzyme 9 -NAD * by means of high performance liquid chromatography. The outer membrane was successfully collected by the methods of Percoll density gradient ultrucentrifugation and ultracentrifugatlon after preferential solubilization of the membrane with Mgt+ and Triton X-100. The enzyme was purified by the series of procedures including extraction of outer membrane protein with EDTA and lysozyme, and fractionation by column chromatography on QAE-Sephadex Q-50, and subsequently Sephadex G-100. The enzyme showed its optimal activity at 85 °C, pH 9.5, and in the presence of 1.5% (vol/vol) Triton X-100. The enzyme exhibited a native molecular size of $8,000 by nondenaturing polyacrylamide gel electrophoresis and bad an apparent Km of 4.72 mM for 2-furaldehyde.
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