Growth of Rhodococcus sp. strain BCP1 on gaseous n-alkanes: new metabolic insights and transcriptional analysis of two soluble di-iron monooxygenase genes.

Martina Cappelletti,Davide Zannoni,Stefano Fedi,Raymond J Turner,Giorgio Milazzo,Alessandro Presentato,Dario Frascari

doi:10.3389/fmicb.2015.00393

Martina Cappelletti, Davide Zannoni + Show 5 more

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https://doi.org/10.3389/fmicb.2015.00393

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Abstract

Rhodococcus sp. strain BCP1 was initially isolated for its ability to grow on gaseous n-alkanes, which act as inducers for the co-metabolic degradation of low-chlorinated compounds. Here, both molecular and metabolic features of BCP1 cells grown on gaseous and short-chain n-alkanes (up to n-heptane) were examined in detail. We show that propane metabolism generated terminal and sub-terminal oxidation products such as 1- and 2-propanol, whereas 1-butanol was the only terminal oxidation product detected from n-butane metabolism. Two gene clusters, prmABCD and smoABCD—coding for Soluble Di-Iron Monooxgenases (SDIMOs) involved in gaseous n-alkanes oxidation—were detected in the BCP1 genome. By means of Reverse Transcriptase-quantitative PCR (RT-qPCR) analysis, a set of substrates inducing the expression of the sdimo genes in BCP1 were assessed as well as their transcriptional repression in the presence of sugars, organic acids, or during the cell growth on rich medium (Luria–Bertani broth). The transcriptional start sites of both the sdimo gene clusters were identified by means of primer extension experiments. Finally, proteomic studies revealed changes in the protein pattern induced by growth on gaseous- (n-butane) and/or liquid (n-hexane) short-chain n-alkanes as compared to growth on succinate. Among the differently expressed protein spots, two chaperonins and an isocytrate lyase were identified along with oxidoreductases involved in oxidation reactions downstream of the initial monooxygenase reaction step.

Highlights

Gaseous n-alkanes ranging from ethane (C2) to n-butane (C4) are categorized as non-methane hydrocarbons
Our present knowledge on the metabolism of short-chain n-alkanes is based on studies in Gram-negative bacteria, when the most common bacterial species isolated from natural habitats that utilize gaseous n-alkanes are Gram-positive strains belonging to the CMNR group
BCP1 Growth on Short-Chain n-Alkanes and on Putative Metabolic Intermediates It has previously been reported that Rhodococcus sp. strain BCP1 has the capacity to grow on n-alkanes (Frascari et al, 2006; Cappelletti et al, 2011)

Summary

Introduction

Gaseous n-alkanes ranging from ethane (C2) to n-butane (C4) are categorized as non-methane hydrocarbons. Volatile aliphatic n-alkanes comprise of gaseous and liquid n-alkanes up to BCP1 growth on short n-alkanes n-heptane (C7). Increasing concentrations of these n-alkanes in the atmosphere is problematic because of their contributions to ozone enhancement and photochemical smog formation (Chan et al, 2006; Shennan, 2006). Aerobic bacterial metabolism of gaseous- and short-chain liquid n-alkanes has received little attention compared to that of methane (C1) and/or longer chain n-alkanes (C >12) (Kotani et al, 2006). Our present knowledge on the metabolism of short-chain n-alkanes is based on studies in Gram-negative bacteria, when the most common bacterial species isolated from natural habitats that utilize gaseous n-alkanes are Gram-positive strains belonging to the CMNR group (including Corynebacterium, Nocardia, Mycobacterium, and Rhodococcus genera). Thauera butanivorans (formerly called “Pseudomonas butanovora”) is the best characterized Gram-negative microorganism able to metabolize gaseous n-alkanes, while the majority of data on gaseous n-alkane microbial growth is based upon the microbial ability to utilize putative metabolic intermediates such as propanal, acetone, and acetal (Ashraf et al, 1994; Arp, 1999; Kulikova and Bezborodov, 2001; Sluis et al, 2002; Kotani et al, 2003; Dubbels et al, 2007; Cooley et al, 2009)

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