3-Hydroxybenzoate 6-Hydroxylase from Rhodococcus jostii RHA1 Contains a Phosphatidylinositol Cofactor

3-Hydroxybenzoate 6-hydroxylase (3HB6H, EC 1.13.14.26) is a FAD-dependent monooxygenase involved in the catabolism of aromatic compounds in soil microorganisms. 3HB6H is unique among flavoprotein hydroxylases in that it harbors a phospholipid ligand. The purified protein obtained from expressing the...

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Published in:Frontiers in microbiology 2017-06, Vol.8, p.1110-1110
Main Authors: Montersino, Stefania, Te Poele, Evelien, Orru, Roberto, Westphal, Adrie H, Barendregt, Arjan, Heck, Albert J R, van der Geize, Robert, Dijkhuizen, Lubbert, Mattevi, Andrea, van Berkel, Willem J H
Format: Article
Language:English
Subjects:
Expression strain
Flavoprotein
Microbiology
Monooxygenase
Phospholipid
Rhodococcus
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Summary:3-Hydroxybenzoate 6-hydroxylase (3HB6H, EC 1.13.14.26) is a FAD-dependent monooxygenase involved in the catabolism of aromatic compounds in soil microorganisms. 3HB6H is unique among flavoprotein hydroxylases in that it harbors a phospholipid ligand. The purified protein obtained from expressing the gene encoding 3HB6H from RHA1 in the host contains a mixture of phosphatidylglycerol and phosphatidylethanolamine, which are the major constituents of cytoplasmic membrane. Here, we purified 3HB6H ( HB6H) produced in the host RHA#2 by employing a newly developed actinomycete expression system. Biochemical and biophysical analysis revealed that 3HB6H possesses similar catalytic and structural features as 3HB6H, but now contains phosphatidylinositol, which is a specific constituent of actinomycete membranes. Native mass spectrometry suggests that the lipid cofactor stabilizes monomer-monomer contact. Lipid analysis of 3HB6H from NCIMB 9867 ( 3HB6H) produced in supports the conclusion that 3HB6H enzymes have an intrinsic ability to bind phospholipids with different specificity, reflecting the membrane composition of their bacterial host.
ISSN:1664-302X
1664-302X
DOI:10.3389/fmicb.2017.01110