Project description:A metagenomic library of sea sediment metagenome containing 245,000 recombinant clones representing ~ 2.45 Gb of sea sediment microbial DNA was constructed. Two unique arsenic resistance clones, A7 and A12, were identified by selection on sodium arsenite containing medium. Clone A7 showed a six-fold higher resistance to arsenate [As(V)], a three-fold higher resistance to arsenite [As(III)] and significantly increased resistance to antimony [Sb(III)], while clone A12 showed increased resistance only to sodium arsenite and not to the other two metalloids. The clones harbored inserts of 8.848 Kb and 6.771 Kb, respectively. Both the clones possess A + T rich nucleotide sequence with similarity to sequences from marine psychrophilic bacteria. Sequence and transposon-mutagenesis based analysis revealed the presence of a putative arsenate reductase (ArsC), a putative arsenite efflux pump (ArsB/ACR) and a putative NADPH-dependent FMN reductase (ArsH) in both the clones and also a putative transcriptional regulatory protein (ArsR) in pA7. The increased resistance of clone A7 to As(V), As(III) and Sb(III) indicates functional expression of ArsC and ArsB proteins from pA7. The absence of increased As(V) resistance in clone A12 may be due to the expression of a possible inactive ArsC, as conserved Arg60 residue in this protein was replaced by Glu60, while the absence of Sb(III) resistance may be due to the presence of an ACR3p-type arsenite pump, which is known to lack antimony transport ability.
Project description:We present a comprehensive data set that describes an anaerobic microbial consortium native to polychlorinated biphenyl (PCB)-contaminated sediments. Obtained from sediment microcosms incubated for 200 days, the data set includes 4 metagenomes, 4 metatranscriptomes (in duplicate), and 62 metagenome-assembled genomes and captures microbial community interactions, structure, and function relevant to anaerobic PCB biodegradation.
Project description:Here, we describe the metagenome and functional composition of a microbial community in a historically metal-contaminated tropical freshwater stream sediment. The sediment was collected from the Mina Stream located in the Iron Quadrangle (Brazil), one of the world's largest mining regions. Environmental DNA was extracted and was sequenced using SOLiD technology, and a total of 7.9 Gbp was produced. A taxonomic profile that was obtained by comparison to the Greengenes database revealed a complex microbial community with a dominance of Proteobacteria and Parvarcheota. Contigs were recruited by bacterial and archaeal genomes, especially Candidatus Nitrospira defluvii and Nitrosopumilus maritimus, and their presence implicated them in the process of N cycling in the Mina Stream sediment (MSS). Functional reconstruction revealed a large, diverse set of genes for ammonium assimilation and ammonification. These processes have been implicated in the maintenance of the N cycle and the health of the sediment. SEED subsystems functional annotation unveiled a high degree of diversity of metal resistance genes, suggesting that the prokaryotic community is adapted to metal contamination. Furthermore, a high metabolic diversity was detected in the MSS, suggesting that the historical arsenic contamination is no longer affecting the prokaryotic community. These results expand the current knowledge of the microbial taxonomic and functional composition of tropical metal-contaminated freshwater sediments.
Project description:We sequenced two metagenomes from upper sediment layers (0 to 5 and 6 to 10 cm) from the Kanawha River, West Virginia. The watershed includes inputs from the forested Appalachian Mountains, surface coal mining, municipal residues, and extensive chemical manufacturing. The dominant bacterial phyla were Proteobacteria, Bacteriodetes, Firmicutes, Actinobacteria, and Chloroflexi Xenobiotic degradation pathways were present.
Project description:L-Carnosine is a natural biologically active dipeptide with critical physiological functions, such as antioxidant, antiglycation, and cytoplasmic buffering properties. Direct enzymatic synthesis is a promising way for L-carnosine production. In this study, a new aminopeptidase (gene_236976) with synthetic activity toward L-carnosine was identified by a metagenome mining approach from deep-sea sediment and functionally expressed in Escherichia coli. The enzyme shared a low identity of 14.3% with reported L-carnosine dipeptidase (SmPepD) from Serratia marcescens. β-Alanine methyl ester was proven to be the best substrate for the synthesis, and no ATP was needed for the enzymatic reaction. The enzyme activity was increased by structure-guided rational design. Only the mutant of G310 site gave positive results, and G310A mutant showed the best performance among the site-direct saturation mutagenesis, indicating that the additional CH3 group of mutant G310A was the main factor affecting the enzymatic activity. The engineered enzyme produced about 10 mM L-carnosine was produced from substrates of 50 mM β-alanine methyl ester and 50 mM L-histidine, under a tentatively optimized condition. This study enriched the enzyme resources for developing the microbial synthesis process of L-carnosine production.