Publications

• Discovery of two novel oxidases using a high-throughput activity screen.

  Rembeza E., Boverio A., Fraaije M.W., Engqvist M.

  Discovery of novel enzymes is a challenging task, yet a crucial one, due to their increasing relevance as chemical catalysts and biotechnological tools. In our work we present a high-throughput screening approach to discovering novel activities. A screen of 96 putative oxidases with 23 substrates ... >> More

  Discovery of novel enzymes is a challenging task, yet a crucial one, due to their increasing relevance as chemical catalysts and biotechnological tools. In our work we present a high-throughput screening approach to discovering novel activities. A screen of 96 putative oxidases with 23 substrates led to the discovery of two new enzymes. The first enzyme, N-acetyl-D-hexosamine oxidase (EC 1.1.3.29) from Ralstonia solanacearum, is a vanillyl alcohol oxidase-like flavoprotein displaying the highest activity with N-acetylglucosamine and N-acetylgalactosamine. Before our discovery of the enzyme, its activity was an orphan one - experimentally characterized but lacking the link to amino acid sequence. The second enzyme, from an uncultured marine euryarchaeota, is a long-chain alcohol oxidase (LCAO, EC 1.1.3.20) active with a range of fatty alcohols, with 1-dodecanol being the preferred substrate. The enzyme displays no sequence similarity to previously characterised LCAOs, and thus is a completely novel representative of a protein with such activity. << Less

  ChemBioChem 0:0-0(2021) [PubMed] [EuropePMC]

  This publication is cited by 6 other entries.

• Cloning and characterization of long-chain fatty alcohol oxidase LjFAO1 in lotus japonicus.

  Zhao S., Lin Z., Ma W., Luo D., Cheng Q.

  The Lotus japonicus EST database was searched against Arabidopsis thaliana AtFAO3, 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 ... >> More

  The Lotus japonicus EST database was searched against Arabidopsis thaliana AtFAO3, 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 LjFAO1 genomic DNA was amplified by PCR, to give a product 3.6 kb in length. Comparison between the LjFAO1 cDNA and genomic DNA revealed that the LjFAO1 contains 3 exons and 2 introns. RT-PCR analysis showed that the LjFAO1 was expressed in the whole plant, with the highest expression level in the apex and the lowest expression level in the siliques. The LjFAO1 gene 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 LjFAO1 exhibited 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 Km values 59.6 +/- 14.8 (microM), 40.9 +/-8.2 (microM) and 19.4 +/-1.5 (microM), respectively, suggesting apparent differences in substrate preferences with AtFAO3. << Less

  Biotechnol. Prog. 24:773-779(2008) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.