title: Community proteomics provides functional insight into polyhydroxyalkanoate production by a mixed microbial culture cultivated on fermented dairy manure.
abstract: 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.
authors.affiliation: Department of Biological Sciences, University of Idaho, Moscow, ID, 83844-3051, USA.
authors.forename: Andrea J
authors.initials: AJ
authors.lastname: Hanson
authors.affiliation: Department of Civil Engineering, University of Idaho, 875 Perimeter Dr. MS1022, Moscow, ID, 83844-1022, USA.
authors.forename: Nicholas M
authors.initials: NM
authors.lastname: Guho
authors.affiliation: Food Research Center, University of Idaho and Washington State University School of Food Science, Moscow, ID, 83844-1052, USA.
authors.forename: Andrzej J
authors.initials: AJ
authors.lastname: Paszczynski
authors.affiliation: Department of Civil Engineering, University of Idaho, 875 Perimeter Dr. MS1022, Moscow, ID, 83844-1022, USA. ecoats@uidaho.edu.
authors.forename: Erik R
authors.initials: ER
authors.lastname: Coats
mesh_terms: D000332:Aerobiosis; D001419:Bacteria / Q000737:chemistry / Q000145:classification / Q000235:genetics; D061249:Batch Cell Culture Techniques; D019149:Bioreactors / Q000382:microbiology*; D058448:Biota; D002853:Chromatography, Liquid; D004275:DNA, Ribosomal / Q000737:chemistry / Q000235:genetics; D015180:Electrophoresis, Gel, Two-Dimensional; D005285:Fermentation; D008372:Manure / Q000382:microbiology*; D054813:Polyhydroxyalkanoates / Q000378:metabolism*; D020543:Proteome / Q000032:analysis*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D017422:Sequence Analysis, DNA; D053719:Tandem Mass Spectrometry
chemical_list: D004275:DNA, Ribosomal; D008372:Manure; D054813:Polyhydroxyalkanoates; D020543:Proteome; D012336:RNA, Ribosomal, 16S
keywords: Aerobic dynamic feeding (ADF); Feast-famine response; Gel-based microbial community proteomics; Illumina sequencing; LC-MS/MS; Polyhydroxyalkanoates (PHAs); Volatile fatty acids (VFAs)