PXD048763

PXD048763 is an original dataset announced via ProteomeXchange.

Title	Bacterial stigmasterol degradation involving radical flavin delta-24 desaturase and molybdenum-dependent C26 hydroxylase
Description	Sterols are ubiquitous membrane constituents that persist to a large extent in the environment due to their water insolubility and chemical inertness. Recently, an oxygenase-independent sterol degradation pathway was discovered in a cholesterol-grown denitrifying bacterium Sterolibacterium (S.) denitrificans. It achieves hydroxylation of the unactivated primary C26 of the isoprenoid side chain to an allylic alcohol via a phosphorylated intermediate in a four-step ATP-dependent enzyme cascade. However, this pathway is incompatible with the degradation of widely distributed steroids containing a double bond at C-22 in the isoprenoid side chain such as the plant sterol stigmasterol. Here, we have enriched a prototypical delta-24 desaturase from S. denitrificans, which catalyses the electron acceptor-dependent oxidation of the intermediate stigmast-1,4-diene-3-one (SDO) to a conjugated 22,(E)24-diene. We suggest an 44 architecture of the 440 kDa enzyme, with each subunit covalently binding an FMN cofactor to a histidyl residue. As isolated, both flavins are present as red semiquinone radicals, which can be reduced by SDO but cannot be oxidised even with strong oxidising agents. We propose a mechanism involving an allylic radical intermediate in which two flavin semiquinones each abstract one hydrogen atom from the substrate. The conjugated delta-22,24 moiety formed allows for the subsequent hydroxylation of the terminal C26 with water by a heterologously produced molybdenum-dependent steroid C26 dehydrogenase. In conclusion, the pathway elucidated for delta-22 steroids achieves oxygen-independent hydroxylation of the isoprenoid side chain by bypassing the ATP-dependent formation of a phosphorylated intermediate.
HostingRepository	PRIDE
AnnounceDate	2024-10-22
AnnouncementXML	Submission_2024-10-22_06:39:50.041.xml
DigitalObjectIdentifier	https://dx.doi.org/10.6019/PXD048763
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	Friedel Drepper
SpeciesList	scientific name: Sterolibacterium denitrificans; NCBI TaxID: 157592;
ModificationList	1'-(8alpha-FMN)-L-histidine
Instrument	Q Exactive Plus

Revision	Datetime	Status	ChangeLog Entry
0	2024-01-22 06:17:04	ID requested
1	2024-05-21 06:53:24	announced
⏵ 2	2024-10-22 06:39:54	announced	2024-10-22: Updated project metadata.

10.1016/j.jbc.2024.107243;

10.6019/PXD048763;

Zhan T, Jacoby C, Jede M, Knapp B, Ferlaino S, G, ü, nter A, Drepper F, M, ü, ller M, Weber S, Boll M, Bacterial stigmasterol degradation involving radical flavin delta-24 desaturase and molybdenum-dependent C26 hydroxylase. J Biol Chem, 300(5):107243(2024) [pubmed]

submitter keyword: anaerobic metabolism, molybdenum enzyme,steroid degradation, flavin radical enzyme, stigmasterol, desaturasem

Friedel Drepper
contact affiliation	University of Freiburg, Institute of Biology II, Schänzlestr. 1, 79104 Freiburg, Germany
contact email	f.drepper@biologie.uni-freiburg.de
lab head
Friedel Drepper
contact affiliation	University of Freiburg
contact email	f.drepper@biologie.uni-freiburg.de
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD048763
     1. Label: PRIDE project
     2. Name: Bacterial stigmasterol degradation involving radical flavin delta-24 desaturase and molybdenum-dependent C26 hydroxylase