Community proteomics provides functional insight into polyhydroxyalkanoate production by a mixed microbial culture cultivated on fermented dairy manure
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Polyhydroxyalkanoates (PHAs) are bio-based, biodegradable polyesters that can be produced from organic-rich waste streams using mixed microbial cultures (MMCs). To maximize PHA production, MMCs are enriched for bacteria with a high polymer storage capacity through the application of aerobic dynamic feeding (ADF) in a sequencing batch reactor (SBR), which consequently induces a feast-famine metabolic response. Though the feast-famine response is generally understood empirically at a macro-level, the molecular level is less refined. The objective of this study was to investigate the microbial community composition and proteome profile of an enriched MMC cultivated on fermented dairy manure. The enriched MMC exhibited a feast-famine response and was capable of producing up to 40 % (wt. basis) PHA in a fed-batch reactor. High-throughput 16S rRNA gene sequencing revealed a microbial community dominated by Meganema, a known PHA-producing genus not often observed in high abundance in enrichment SBRs. The application of the proteomic methods two-dimensional electrophoresis and LC-MS/MS revealed PHA synthesis, energy generation, and protein synthesis prominently occurring during the feast phase, corroborating bulk solution variable observations and theoretical expectations. During the famine phase, nutrient transport, acyl-CoA metabolism, additional energy generation, and housekeeping functions were more pronounced, informing previously under-determined MMC functionality under famine conditions. During fed-batch PHA production, acetyl-CoA acetyltransferase and PHA granule-bound phasin proteins were in increased abundance relative to the SBR, supporting the higher PHA content observed. Collectively, the results provide unique microbial community structural and functional insight into feast-famine PHA production from waste feedstocks using MMCs.
KeywordsAerobic dynamic feeding (ADF) Feast-famine response Polyhydroxyalkanoates (PHAs) Volatile fatty acids (VFAs) Gel-based microbial community proteomics LC-MS/MS Illumina sequencing
The authors acknowledge Dr. Armando McDonald at the University of Idaho for the use of GC-MS. The authors acknowledge Mr. Dan New and Dr. Alida Gerritsen at the Institute for Bioinformatics and Evolutionary Studies Genomics Resources Core at the University of Idaho, and Dr. Matt Settles at the Bioinformatics Core at the Genome Center at the University of California-Davis for the technical assistance related to the 16S rRNA gene sequencing and bioinformatics analysis. The authors acknowledge Dr. Lee Deobald at the Mass Spectrometry Core at the University of Idaho for the technical assistance related to the LC-MS/MS proteomic analysis.
Compliance with ethical standards
This material is based on work supported by the National Science Foundation under Grant Number CBET-0950498, the Environmental Protection Agency Science to Achieve Results Fellowship Program, and the National Institute of General Medical Sciences from the National Institutes of Health P30 GM103324. Any opinions, findings, and conclusions or recommendations expressed in this material are those of the authors and do not necessarily reflect the views of the funding agency.
Conflict of interest
The authors declare that they have no conflicts of interest.
This article does not contain any studies with human participants or animals performed by any of the authors.
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