Sphingobiumsp. SYK-6 syringateO-demethylase gene is regulated by DesX, unlike other vanillate and syringate catabolic genes regulated by DesR

Abstract

Syringate and vanillate are the major metabolites of lignin biodegradation. In Sphingobium sp. strain SYK-6, syringate is O demethylated to gallate by consecutive reactions catalyzed by DesA and LigM, and vanillate is O demethylated to protocatechuate by a reaction catalyzed by LigM. The gallate ring is cleaved by DesB, and protocatechuate is catabolized via the protocatechuate 4,5-cleavage pathway. The transcriptions of desA, ligM, and desB are induced by syringate and vanillate, while that of ligM and desB are negatively regulated by the MarR-type transcriptional regulator DesR, which is not involved in desA regulation. Here we clarified the regulatory system for desA transcription by analyzing the IclR-type transcriptional regulator desX, located downstream of desA. Quantitative reverse transcription (RT)-PCR analyses of a desX mutant indicates that the transcription of desA was negatively regulated by DesX. In contrast, DesX was not involved in the regulation of ligM and desB. The ferulate catabolic genes (ferBA) under the control of a MarR-type transcriptional regulator FerC are located upstream of desA. RT-PCR analyses suggest that the ferB-ferA-SLG_25010-desA gene cluster consists of the ferBA operon and the SLG_25010-desA operon. Promoter assays reveal that a syringate- and vanillate-inducible promoter is located upstream of SLG_25010. Purified DesX bound to this promoter region, which overlaps with an 18-bp-inverted repeat sequence that appears to be essential for the DNA binding of DesX. Syringate and vanillate inhibited the DNA binding of DesX, indicating that these compounds are effector molecules of DesX.IMPORTANCESyringate is a major degradation product in the microbial and chemical degradation of syringyl lignin. Along with other low-molecular-weight aromatic compounds, syringate is produced by chemical lignin depolymerization. Converting this mixture into value-added chemicals using bacterial metabolism (i.e., biological funneling) is a promising option for lignin valorization. To construct an efficient microbial lignin conversion system, it is necessary to identify and characterize the genes involved in the uptake and catabolism of lignin-derived aromatic compounds and elucidate their transcriptional regulation. In this study, we found that the transcription of desA, encoding syringate O-demethylase in SYK-6, is regulated by an IclR-type of transcriptional regulator, DesX. The findings of this study, combined with our previous results on desR (a MarR transcriptional regulator that controls the transcription of ligM and desB), provide an overall picture of the transcriptional regulatory systems for syringate and vanillate catabolism in SYK-6.

Bacterial catabolic system of acetovanillone and acetosyringone useful for upgrading aromatic compounds obtained through chemical lignin depolymerization

“…Intermediate metabolites generated during SYK-6 incubation with AV were identified to determine the catabolic pathway of AV in SYK-6. SYK-6 cells grown with AV were incubated with 1 mM AV in Wx minimal medium (48) for 33 h, and high-performance liquid chromatography (HPLC)–mass spectrometry (MS) analyzed the supernatant of the reaction mixture. This analysis indicated that AV disappeared and was converted into compounds I and II with retention times of 2.2 and 2.5 min, respectively (Fig.…”

Bioconversion of Lignocellulosic Biomass into Value Added Products under Anaerobic Conditions: Insight into Proteomic Studies

“…Lignin depolymerization results in different lignin derivatives, and the most common are vainillate and syringate. Subsequently, demethylation of vainillate and syringate by vainillate O-demethylase and syringate O-demethylase, respectively, are important steps to produce protocatechuate and gallate, which as intermediaries enter different pathways for ring cleavage [70,82]. Under anaerobic conditions, protocatechuate and gallate are converted to different key intermediates such as benzoyl-CoA, phloroglucinol, hydroxy-hydroquinone and resorcinol [73].…”