Genome Sequence of the Methanotrophic Poly-β-Hydroxybutyrate Producer Methylocystis parvus OBBP

Abstract

cMethylocystis parvus OBBP is an obligate methylotroph considered the type species of the genus Methylocystis. Two pmoCAB particulate methane monooxygenase operons and one additional singleton pmoC paralog were identified in the sequence. No evidence of genes encoding soluble methane monooxygenase was found. Comparison of M. parvus OBBP and Methylocystis sp. strain Rockwell (ATCC 49242) suggests that both species should be taxonomically classified in different genera. Methane is a greenhouse gas that accumulates in the atmosphere and contributes to climate change and global warming (7). Methane-oxidizing bacteria (methanotrophs), a subset of methylotrophs, are a group of bacteria with a specialized metabolism able to utilize C 1 compounds (2, 5). Methanotrophs are essential for the global methane cycle since they are capable of oxidizing methane (or methanol) to formaldehyde, which can be transformed into biomass or oxidized to carbon dioxide (3, 4). Type II methanotrophs belong to Alphaproteobacteria. There are four genera, Methylosinus, Methylocystis, Methylocella, and Methylocapsa (13). Methylocystis parvus OBBP (NCIMB 11129) is an obligate methylotroph originally isolated as a methane-utilizing bacterium and is considered the type species of the genus Methylocystis (16). This strain has industrial interest because it is able to produce poly-␤-hydroxybutyrate (PHB) from methane (9).The genome of M. parvus OBBP was sequenced using the Titanium kit and GS-FLX pyrosequencing equipment from Roche. Preliminary assembly of raw reads was performed using Newbler software from Roche. In order to improve the assembly quality in-house, Perl shell scripts were designed. A quality draft of 108 contigs was obtained. The genome was structurally and functionally annotated using RAST (1), an automated genome annotation system, and functions, names, and general properties of the gene products were predicted using this method. The M. parvus OBBP genome is 4.5 Mbp in size, contains 63.4% GC, encodes 49 RNAs, and contains 4,594 coding sequences.Two pmoCAB particulate methane monooxygenase operons (8, 12) and one additional singleton pmoC paralog were identified in the sequence, in contrast with the genome of Methylocystis sp. strain Rockwell (ATCC 49242), where only a single pmoCAB particulate methane monooxygenase operon and four singleton pmoC paralogs were recognized (15). Genes encoding soluble methane monooxygenase were absent in both cases.Other proteins involved in methane oxidation were identified; these include a methanol dehydrogenase, a set of enzymes that are involved in pyrroloquinoline quinone synthesis and tetrahydrofolate-and tetrahydromethanopterin-linked pathways, and NAD-linked formate dehydrogenase. Enzymes required for the serine cycle were identified, confirming that M. parvus OBBP possess a methane assimilation pathway linked to the central metabolite pathways via this cycle (6).Enzymes involved in the ethylmalonyl-coenzyme A (ethylmalonyl-CoA) pathway for glyoxylate regeneration were detected in M. parvus ...

Genome mining for methanobactins

“…Two Methylosinus species ( Methylosinus sp. LW3 and Methylocystis parvus OBBP, which may be misclassified as Methylocystis [40]) have two distinct MbnA genes encoding unique Mbns. While it is not uncommon for bacteria to produce multiple siderophores to control iron acquisition in different environments [41], a similar phenomenon has not yet been observed for chalkophores.…”

“…Of the methanotroph species, operons are present in both strains that undergo the copper switch from sMMO to pMMO (for example, Methylosinus trichosporium OB3b [28], Methylocystis str. M [80,81], Methylocystis hirsuta CSC-1 [82]) and those that only express pMMO (for example, Methylocystis parvus OBBP [40], Methylocystis rosea SV97T [83]). The 16 species are not limited to methanotrophic bacteria, providing compelling evidence that Mbn-like compounds may play a broader role in proteobacterial metal homeostasis.…”

“…SC2 (GenBank: NC_018485) [43], Methylobacterium sp. B34 (GenBank: BADE01000957), Methylocystis parvus OBBP (GenBank: AJTV01000003 and AJTV01000041) [40] , Methylocystis rosea SV97T (IMG: A3OODRAFT_scaffold1.1) , Methylosinus sp. LW3 (IMG: MetLW3DRAFT_contig2.2), Methylosinus sp.…”

Genome Analysis Coupled with Physiological Studies Reveals a Diverse Nitrogen Metabolism in Methylocystis sp. Strain SC2

“…These include two PHB depolymerases (BN69_2992 and BN69_3262), one polyhydroxyalkonate synthesis repressor (PhbR [BN69_3069]), an acetyl-CoA acetyltransferase (PhbA [BN69_3068]), one acetoacetyl-CoA reductase (PhbB [BN69_3067]), and two phasin homologs (BN69_0212, BN69_1107). The phbR , phbA , and phbB genes form a cluster, but in an orientation different from that in the PHB-producing methanotroph Methylocystis parvus OBB ( phbABR ) [40]. The use of PHB as a reducing power for denitrification has been shown in microbial granules in bioreactors [97].…”

“…At present, twelve methanotroph genomes are available in public databases and more are being sequenced. The available sequences include those of the alphaproteobacterial methanotrophs Methylosinus trichosporium OB3b [39], Methylocystis parvus OBBP [40], Methylocystis sp. strain Rockwell [41], Methylocystis sp.…”