The Glycobiome of the Rumen Bacterium Butyrivibrio proteoclasticus B316T Highlights Adaptation to a Polysaccharide-Rich Environment

Abstract

Determining the role of rumen microbes and their enzymes in plant polysaccharide breakdown is fundamental to understanding digestion and maximising productivity in ruminant animals. Butyrivibrio proteoclasticus B316T is a Gram-positive, butyrate-forming rumen bacterium with a key role in plant polysaccharide degradation. The 4.4Mb genome consists of 4 replicons; a chromosome, a chromid and two megaplasmids. The chromid is the smallest reported for all bacteria, and the first identified from the phylum Firmicutes. B316 devotes a large proportion of its genome to the breakdown and reassembly of complex polysaccharides and has a highly developed glycobiome when compared to other sequenced bacteria. The secretion of a range of polysaccharide-degrading enzymes which initiate the breakdown of pectin, starch and xylan, a subtilisin family protease active against plant proteins, and diverse intracellular enzymes to break down oligosaccharides constitute the degradative capability of this organism. A prominent feature of the genome is the presence of multiple gene clusters predicted to be involved in polysaccharide biosynthesis. Metabolic reconstruction reveals the absence of an identifiable gene for enolase, a conserved enzyme of the glycolytic pathway. To our knowledge this is the first report of an organism lacking an enolase. Our analysis of the B316 genome shows how one organism can contribute to the multi-organism complex that rapidly breaks down plant material in the rumen. It can be concluded that B316, and similar organisms with broad polysaccharide-degrading capability, are well suited to being early colonizers and degraders of plant polysaccharides in the rumen environment.

Carbohydrate transporting membrane proteins of the rumen bacterium, Butyrivibrio proteoclasticus

“…This gives rise to a model of plant cell wall breakdown in which the secreted enzymes generate a variety of complex oligosaccharides which are transported into the cell for further metabolism. The clustering of genes encoding intracellular polysaccharide degrading enzymes with genes for transporters, transcriptional regulators and environmental sensors in several polysaccharide utilisation loci [1] lends support to this model. The transport of plant derived sugars or oligosaccharides into the cell is hypothesised to be by specific membrane bound proteins, including more than 20 predicted ATP-binding cassette (ABC) transporter systems (TC3.A.1 in the Transporter Classification Database) [2].…”

“…Open reading frames (ORFs) were generated as reported by Kelly et al [1] and theoretical pI and Mr calculated for each locus. The prediction of membrane proteins was based on the amino acid distribution of the protein sequence [8].…”

“…The genome of this strain has been sequenced [1] and found to encode a large complement of polysaccharide degrading enzymes. The enzymes that initiate polysaccharide breakdown are secreted by B. proteoclasticus but the majority of the enzymes involved in carbohydrate metabolism are predicted to be intracellular.…”

A 3-D Model of a Perennial Ryegrass Primary Cell Wall and Its Enzymatic Degradation

“…For example, F. succinogenes appears to readily utilise glucose and cellobiose [72], whereas some microorganisms utilise larger oligosaccharides more effectively [73]. Certainly a study of Butyrivibrio proteoclasticus B316 T enzymes suggests a variety of complex oligosaccharides resulting from extracellular hydrolysis are metabolized within the cell [74]. Thus the model outputs could be used as inputs into models of microbial growth to make predictions of the competitiveness of different microbial species for different compositions of the PCW.…”

Bacterial Typing and Identification By Genomic Analysis of 16S–23S rRNA Intergenic Transcribed Spacer (ITS) Sequences

“…In most bacteria, the 16S, 23S and 5S rRNA genes are all present in the rrn operons, in a single copy per operon, with approximate lengths highly conserved within genomes and species but varying more between species. The variations in intergenic spacers between the 16S and 23S rRNA genes (ITS1) have been grouped according to the type of tRNA gene they contain, with examples of species that have operons with these types of ITS sequences summarised in Gurtler (1999) and further examples for each type given shown in the diagram as follows: (A) C. difficile (Gurtler & Grando, 2013); (B) and (C) Staphylococcus aureus (Gurtler & Barrie, 1995); (C) and (D) Escherichia coli (Condon, Squires, & Squires, 1995) and Salmonella typhimurium (Christensen, Moller, Vogensen, & Olsen, 2000;Perez Luz, Rodriguez-Valera, Lan, & Reeves, 1998); (D)-(G) Vibrio species (Chun, Huq, & Colwell, 1999;GonzalezEscalona, Romero, Guzman, & Espejo, 2006); and (H)-(J) Butyrivibrio proteoclasticus (Kelly et al, 2010;Li et al, 2014). The line without tRNA gene boxes at the top refers to many alleles in many species which do not contain any tRNA genes.…”