A proteomic, reverse-genetic and sequencing approach to reveal genes and enzymes for LAS-surfactant degradation in bacterial communities

Institutionen
  • FB Biologie
Publikationen
    Weiss, Michael; Denger, Karin; Huhn, Thomas; Schleheck, David(2012): Two enzymes of a complete degradation pathway for linear alkylbenzenesulfonate (LAS) surfactants : 4-sulfoacetophenone Baeyer-Villiger monooxygenase and 4-sulfophenylacetate esterase in Comamonas testosteroni KF-1 Applied and Environmental Microbiology ; 78 (2012), 23. - S. 8254-8263. - ISSN 0099-2240. - eISSN 1098-5336

Two enzymes of a complete degradation pathway for linear alkylbenzenesulfonate (LAS) surfactants : 4-sulfoacetophenone Baeyer-Villiger monooxygenase and 4-sulfophenylacetate esterase in Comamonas testosteroni KF-1

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Complete biodegradation of the surfactant linear alkylbenzenesulfonate (LAS) is accomplished by complex bacterial communities in two steps. First, all LAS congeners are degraded into about 50 sulfophenylcarboxylates (SPC), one of which is 3-(4-sulfophenyl)butyrate (3-C(4)-SPC). Second, these SPCs are mineralized. 3-C(4)-SPC is mineralized by Comamonas testosteroni KF-1 in a process involving 4-sulfoacetophenone (SAP) as a metabolite and an unknown inducible Baeyer-Villiger monooxygenase (BVMO) to yield 4-sulfophenyl acetate (SPAc) from SAP (SAPMO enzyme); hydrolysis of SPAc to 4-sulfophenol and acetate is catalyzed by an unknown inducible esterase (SPAc esterase). Transcriptional analysis showed that one of four candidate genes for BVMOs in the genome of strain KF-1, as well as an SPAc esterase candidate gene directly upstream, was inducibly transcribed during growth with 3-C(4)-SPC. The same genes were identified by enzyme purification and peptide fingerprinting-mass spectrometry when SAPMO was enriched and SPAc esterase purified to homogeneity by protein chromatography. Heterologously overproduced pure SAPMO converted SAP to SPAc and was active with phenylacetone and 4-hydroxyacetophenone but not with cyclohexanone and progesterone. SAPMO showed the highest sequence homology to the archetypal phenylacetone BVMO (57%), followed by steroid BVMO (55%) and 4-hydroxyacetophenone BVMO (30%). Finally, the two pure enzymes added sequentially, SAPMO with NADPH and SAP, and then SPAc esterase, catalyzed the conversion of SAP via SPAc to 4-sulfophenol and acetate in a 1:1:1:1 molar ratio. Hence, the first two enzymes of a complete LAS degradation pathway were identified, giving evidence for the recruitment of members of the very versatile type I BVMO and carboxylester hydrolase enzyme families for the utilization of a xenobiotic compound by bacteria.

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Laufzeit: 01.04.2011 – 31.03.2014