Metabolic Regulation as a Consequence of Anaerobic 5-Methylthioadenosine Recycling in Rhodospirillum rubrum.

Justin A North,Robert L Hettich,Jaya Sriram,F Robert Tabita,Karuna Chourey,John A Wildenthal,Ritin Sharma,Christopher D Ecker,Colleen M Cavanaugh

doi:10.1128/mbio.00855-16

Justin A North, Robert L Hettich + Show 7 more

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https://doi.org/10.1128/mbio.00855-16

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Abstract

ABSTRACTRhodospirillum rubrum possesses a novel oxygen-independent, aerobic methionine salvage pathway (MSP) for recycling methionine from 5-methylthioadenosine (MTA), the MTA-isoprenoid shunt. This organism can also metabolize MTA as a sulfur source under anaerobic conditions, suggesting that the MTA-isoprenoid shunt may also function anaerobically as well. In this study, deep proteomics profiling, directed metabolite analysis, and reverse transcriptase quantitative PCR (RT-qPCR) revealed metabolic changes in response to anaerobic growth on MTA versus sulfate as sole sulfur source. The abundance of protein levels associated with methionine transport, cell motility, and chemotaxis increased in the presence of MTA over that in the presence of sulfate. Purine salvage from MTA resulted primarily in hypoxanthine accumulation and a decrease in protein levels involved in GMP-to-AMP conversion to balance purine pools. Acyl coenzyme A (acyl-CoA) metabolic protein levels for lipid metabolism were lower in abundance, whereas poly-β-hydroxybutyrate synthesis and storage were increased nearly 10-fold. The known R. rubrum aerobic MSP was also shown to be upregulated, to function anaerobically, and to recycle MTA. This suggested that other organisms with gene homologues for the MTA-isoprenoid shunt may also possess a functioning anaerobic MSP. In support of our previous findings that ribulose-1,5-carboxylase/oxygenase (RubisCO) is required for an apparently purely anaerobic MSP, RubisCO transcript and protein levels both increased in abundance by over 10-fold in cells grown anaerobically on MTA over those in cells grown on sulfate, resulting in increased intracellular RubisCO activity. These results reveal for the first time global metabolic responses as a consequence of anaerobic MTA metabolism compared to using sulfate as the sulfur source.

Highlights

Rhodospirillum rubrum possesses a novel oxygen-independent, aerobic methionine salvage pathway (MSP) for recycling methionine from 5-methylthioadenosine (MTA), the MTA-isoprenoid shunt
To determine the cellular pathways affected as a consequence of growth on MTA versus sulfate, we compared the proteome profiles of R. rubrum cultures grown in the presence of MTA and those grown in the presence of sulfate as sole sulfur source (Table 1; see Table S1 in the supplemental material)
In eukaryotes and many prokaryotes, MTA is recycled into usable methionine by the universal methionine salvage pathway, which requires oxygen for the final metabolic step to occur (Fig. 1, black arrows)

Summary

Introduction

Rhodospirillum rubrum possesses a novel oxygen-independent, aerobic methionine salvage pathway (MSP) for recycling methionine from 5-methylthioadenosine (MTA), the MTA-isoprenoid shunt This organism can metabolize MTA as a sulfur source under anaerobic conditions, suggesting that the MTA-isoprenoid shunt may function anaerobically as well. In Tetrahymena sp., a multifunctional fusion enzyme catalyzes the dehydratase, enolase/phosphatase, and dioxygenase reactions to generate 2-keto4-methylthiobutyrate (KMTB) from 5-methylthioribulose-1phosphate (MTRu-1P) (Fig. 1, letter L) [11] Some of these variations employ a RubisCO-like protein (RLP) [16,17,18,19,20,21]. In B. subtilis, a Ykr class RLP (Fig. 1, letter G, mtnW) functions as the

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