luxA gene from Enhygromyxa salina encodes a functional homodimeric luciferase

Yudenko, A.; Bazhenov, S. V.; Aleksenko, V. A.; Goncharov, I. M.; Semenov, O.; Remeeva, A.; Nazarenko, V. V.; Kuznetsova, E.; Fomin, V. V.; Konopleva, M. N.; Al Ebrahim, R.; Sluchanko, N. N.; Ryzhykau, Y.; Semenov, Y. S.; Kuklin, A.; Manukhov, I. V.; Gushchin, I.

doi:10.1101/2023.02.27.530324

bioRxiv · 10.1101/2023.02.27.530324

luxA gene from Enhygromyxa salina encodes a functional homodimeric luciferase

Yudenko, A.·Bazhenov, S. V.·Aleksenko, V. A.·Goncharov, I. M.·Semenov, O.·Remeeva, A.·Nazarenko, V. V.·Kuznetsova, E.·Fomin, V. V.·Konopleva, M. N.·Al Ebrahim, R.·Sluchanko, N. N.·Ryzhykau, Y.·Semenov, Y. S.·Kuklin, A.·Manukhov, I. V.·Gushchin, I.

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Abstract

Several clades of luminescent bacteria are known currently. They all contain similar lux operons, which include the genes luxA and luxB encoding a heterodimeric luciferase. The aldehyde oxygenation reaction is catalyzed by the subunit LuxA, while LuxB is inactive. Recently, genomic analysis identified a subset of bacterial species with rearranged lux operons lacking luxB. Here, we show that the product of the luxA gene from the reduced luxACDE operon of Enhygromyxa salina is luminescent upon addition of aldehydes both in vivo in Escherichia coli and in vitro. Overall, EsLuxA is less bright compared with luciferases from Aliivibrio fischeri (AfLuxAB) and Photorhabdus luminescens (PlLuxAB), and most active with medium-chain C4-C9 aldehydes. Crystal structure of EsLuxA determined at the resolution of 2.71 [A] reveals a classical monooxygenase fold, and the protein preferentially forms a dimer in solution. The mobile loop residues 264-293, which form a {beta}-hairpin or a coil in Vibrio harveyi LuxA, form -helices in EsLuxA. Phylogenetic analysis shows EsLuxA and related proteins may be bacterial protoluciferases that arose prior to duplication of the luxA gene and its speciation to luxA and luxB in the previously described luminescent bacteria. Our work paves the way for discovery of new luciferases that have an advantage of being encoded by a single gene.

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Yudenko, A., Bazhenov, S. V., Aleksenko, V. A., Goncharov, I. M., Semenov, O., Remeeva, A., Nazarenko, V. V., Kuznetsova, E., Fomin, V. V., Konopleva, M. N., Al Ebrahim, R., Sluchanko, N. N., Ryzhykau, Y., Semenov, Y. S., Kuklin, A., Manukhov, I. V., Gushchin, I.. 2023-02-27. luxA gene from Enhygromyxa salina encodes a functional homodimeric luciferase. https://doi.org/10.1101/2023.02.27.530324

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