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619261 
Journal Article 
Cloning and Characterization of Long-Chain Fatty Alcohol Oxidase LjFAO1in Lotus japonicus 
Shilan, Z; Zhixin, L; Wei, M; Da, L; Qi, C 
2008 
Yes 
Biotechnology Progress
ISSN: 8756-7938
EISSN: 1520-6033 
24 
3 
773-779 
The Lotus japonicusEST database was searched against Arabidopsis thalianaAtFAO3, a full-length cDNA that encodes a membrane-bound, flavin-containing, hydrogen peroxide generating, long-chain fatty alcohol oxidase. One EST fragment was detected, and the corresponding full-length cDNA was obtained by screening a cDNA library of L. japonicus. The LjFAO1genomic DNA was amplified by PCR, to give a product 3.6 kb in length. Comparison between the LjFAO1cDNA and genomic DNA revealed that the LjFAO1contains 3 exons and 2 introns. RT-PCR analysis showed that the LjFAO1was expressed in the whole plant, with the highest expression level in the apex and the lowest expression level in the siliques. The LjFAO1gene was down-regulated by cold stress in both the apex and the cotelydon of the 8-day old seedlings, the first time that a long-chain alcohol oxidase has been shown to respond directly to stress. The full length cDNA and a C-terminal truncated version were overexpressed in Escherichia coli. The full length version of LjFAO1exhibited long-chain fatty alcohol oxidase activity and was subsequently purified by Ni-NTA chromatography. The active LjFAO1 protein showed substrate specificities toward 1-dodecanol, 1-hexadecanol, and 1,16-hexadecanediol with Kmvalues 59.6 ± 14.8 (M), 40.9 ± 8.2 (M) and 19.4 ± 1.5 (M), respectively, suggesting apparent differences in substrate preferences with AtFAO3. ABSTRACT FROM AUTHOR Copyright of Biotechnology Progress is the property of American Chemical Society and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts) 
ESCHERICHIA coli; CHROMATOGRAPHIC analysis; BIOTECHNOLOGY; CLONING; FATTY alcohols; ARABIDOPSIS thaliana; HYDROGEN peroxide; OXIDASES