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Title: | Characterization of the enzyme involved in the diol synthase metabolic pathway in Pseudomonas aeruginosa |
Author: | Shoja Chaghervand, S Castells, Marc Rabanal Anglada, Francesc Cajal Visa, Yolanda Manresa Presas, Ma. Ángeles (María Ángeles) Estupiñán, Mónica Busquets, Montserrat |
Keywords: | Microscòpia de força atòmica Temperatura Atomic force microscopy Temperature |
Issue Date: | 10-Jun-2022 |
Publisher: | Elsevier |
Abstract: | This study is the first attempt to report the biochemical characterisation of the 10S-dioxygenase (10S-DOX) and 7S, 10S-diol synthase (7,10-DS). Both enzymes showed similar pH profiles with 10S-DOX presenting the highest activity at 30 °C whereas 7,10-DS did not show a clear preferred temperature. These differences were reflected in the thermostability assay, the Km values were 0.89 ± 0.22 mM and 3.26 ± 0.31 mM for 10S-DOX and 7,10-DS, respectively. Inductively coupled plasma mass spectrometry indicated that both enzymes contained bound to haem group Fe2+ as a prosthetic group: 10S-DOX (0.95 mol Fe2+/mol of protein) and 7,10-DS (1.18 mol Fe2+/mol of protein), respectively. Assays using metal cations as cofactors revealed that Mg2+ and Ni2+ enhance 7,10-DS activity, whereas Hg2+ decrease it up to 50 %. The activity of 10S-DOX in the presence of Mn2+ and Fe2+ was reduced to 51.6 % and 61.8. Aggregated proteins producing 10S-DOX and 7,10-DS were characterised as inclusion bodies: IBs-77 and IBs-78 respectively, was performed by Fourier Transform spectroscopy (FT-IR), Atomic Force Microscopy, dye binding, and proteolysis. The specific activity was 1.55 IU/mg for IBs-77 and 1.05 IU/mg for IBs-78. The presence of the oleate-diol synthase pathway in proteobacteria other than Pseudomonas aeruginosa was detected. |
Note: | Versió postprint del document publicat a: https://doi.org/10.1016/j.procbio.2022.017 |
It is part of: | Process Biochemistry, 2022, vol. 120, p. 301-312 |
URI: | http://hdl.handle.net/2445/195944 |
Related resource: | https://doi.org/10.1016/j.procbio.2022.017 |
ISSN: | 1359-5113 |
Appears in Collections: | Articles publicats en revistes (Química Inorgànica i Orgànica) |
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