Expression of 5,8-LDS of Aspergillus fumigatus and its dioxygenase domain: a comparison with 7,8-LDS, 10-dioxygenase, and cyclooxygenase
2011 (English)In: Archives of Biochemistry and Biophysics, ISSN 0003-9861, E-ISSN 1096-0384, Vol. 506, no 2, 216-222 p.Article in journal (Refereed) Published
5,8-Linoleate diol synthase (5,8-LDS) of Aspergillus fumigatus was cloned, expressed, and compared with 7,8-LDS of the Take-all fungus. Replacements of Tyr and Cys in the conserved YRWH and FXXGPHXCLG sequences abolished 8R-dioxygenase (8-DOX) and hydroperoxide isomerase activities, respectively. The predicted α-helices of LDS were aligned with α-helices of cyclooxygenase-1 (COX-1) to identify the 8-DOX domains. N-terminal expression constructs of 5,8- and 7,8-LDS (674 of 1079, and 673 of 1165 residues), containing one additional α-helix compared to cyclooxygenase-1, yielded prominent 8R-DOX activities with apparently unchanged or slightly lower substrate affinities, respectively. Val-328 of 5,8-LDS did not influence the position of oxygenation in contrast to the homologous residues Val-349 of COX-1 and Leu-384 of 10R-dioxygenase. We conclude that ∼675 amino acids are sufficient to support 8-DOX activity.
Place, publisher, year, edition, pages
2011. Vol. 506, no 2, 216-222 p.
Cytochrome P450, fusion protein, heme-dependent peroxidase, hydroperoxide isomerase, LC-MS/MS, oxylipins
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Research subject Biochemical Pharmacology
IdentifiersURN: urn:nbn:se:uu:diva-142985DOI: 10.1016/j.abb.2010.11.022ISI: 000286961600014PubMedID: 21130068OAI: oai:DiVA.org:uu-142985DiVA: diva2:388865