50 research outputs found
Site-directed mutagenesis and stability of the carboxylic acid reductase MAB4714 from Mycobacterium abscessus
Modeling of laser-induced plasmon effects in GNS-DLC-based material for application in X-ray source array sensors
An important direction in the development of X-ray computed tomography sensors in systems with increased scanning speed and spatial resolution is the creation of an array of miniature current sources. In this paper, we describe a new material based on gold nanostars (GNS) embedded in nanoscale diamond-like carbon (DLC) films (thickness of 20 nm) for constructing a pixel current source with photoinduced electron emission. The effect of localized surface plasmon resonance in GNS on optical properties in the wavelength range from UV to near IR, peculiarities of localization of field and thermal sources, generation of high-energy hot electrons, and mechanisms of their transportation in vacuum are investigated. The advantages of the proposed material and the prospects for using X-ray computed tomography in the matrix source are evaluated
One-pot biocatalytic transformation of adipic acid to 6-aminocaproic acid and 1,6-hexamethylenediamine using carboxylic acid reductases and transaminases
Metabolic and evolutionary patterns in the extremely acidophilic archaeon Ferroplasma acidiphilum YT
Ferroplasmaceae represent ubiquitous iron-oxidising extreme acidophiles with a number of unique physiological traits. In a genome-based study of Ferroplasma acidiphilum YT, the only species of the genus Ferroplasma with a validly published name, we assessed its central metabolism and genome stability during a long-term cultivation experiment. Consistently with physiology, the genome analysis points to F. acidiphilum YT having an obligate peptidolytic oligotrophic lifestyle alongside with anaplerotic carbon assimilation. This narrow trophic specialisation abridges the sugar uptake, although all genes for glycolysis and gluconeogenesis, including bifunctional unidirectional fructose 1,6-bisphosphate aldolase/phosphatase, have been identified. Pyruvate and 2-oxoglutarate dehydrogenases are substituted by ‘ancient’ CoA-dependent pyruvate and alpha-ketoglutarate ferredoxin oxidoreductases. In the lab culture, after ~550 generations, the strain exhibited the mutation rate of ≥1.3 × 10−8 single nucleotide substitutions per site per generation, which is among the highest values recorded for unicellular organisms. All but one base substitutions were G:C to A:T, their distribution between coding and non-coding regions and synonymous-to-non-synonymous mutation ratios suggest the neutral drift being a prevalent mode in genome evolution in the lab culture. Mutations in nature seem to occur with lower frequencies, as suggested by a remarkable genomic conservation in F. acidiphilum YT variants from geographically distant populations
Structural and biochemical insights into CRISPR RNA processing by the Cas5c ribonuclease SMU1763 from Streptococcus mutans
Biochemical diversity of carboxyl Esterases and Lipases from Lake Arreo (Spain) : a metagenomic approach
The esterases and lipases from the / hydrolase superfamily exhibit an enormous sequence diversity, fold plasticity, and activities.
Here, we present the comprehensive sequence and biochemical analyses of seven distinct esterases and lipases from the
metagenome of Lake Arreo, an evaporite karstic lake in Spain (42°46=N, 2°59=W; altitude, 655 m). Together with oligonucleotide
usage patterns and BLASTP analysis, our study of esterases/lipases mined from Lake Arreo suggests that its sediment contains
moderately halophilic and cold-adapted proteobacteria containing DNA fragments of distantly related plasmids or chromosomal
genomic islands of plasmid and phage origins. This metagenome encodes esterases/lipases with broad substrate profiles
(tested over a set of 101 structurally diverse esters) and habitat-specific characteristics, as they exhibit maximal activity at alkaline
pH (8.0 to 8.5) and temperature of 16 to 40°C, and they are stimulated (1.5 to 2.2 times) by chloride ions (0.1 to 1.2 M), reflecting
an adaptation to environmental conditions. Our work provides further insights into the potential significance of the
Lake Arreo esterases/lipases for biotechnology processes (i.e., production of enantiomers and sugar esters), because these enzymes
are salt tolerant and are active at low temperatures and against a broad range of substrates. As an example, the ability of a
single protein to hydrolyze triacylglycerols, (non)halogenated alkyl and aryl esters, cinnamoyl and carbohydrate esters, lactones,
and chiral epoxides to a similar extent was demonstrated.The Spanish
Ministry of Economy and Competitiveness (project CSD2007-00005), the
European Community project MAGICPAH (FP7-KBBE-2009-245226),
the European Regional Development Fund (ERDF), and the Government
of Canada through Genome Canada, Ontario Genomics Institute, and
Ontario Research Fund (2009-OGI-ABC-1405 and ORF-GL2-01-004).
M.-E.G. thanks the CSIC for a JAE fellowship.http://aem.asm.org/am201
Identification and characterization of carboxyl esterases of gill chamber-associated microbiota in the deep-sea shrimp rimicaris exoculata by using functional metagenomics
The shrimp Rimicaris exoculata dominates the fauna in deep-sea hydrothermal vent sites along the Mid-Atlantic Ridge (depth,
2,320 m). Here, we identified and biochemically characterized three carboxyl esterases from microbial communities inhabiting
the R. exoculata gill that were isolated by naive screens of a gill chamber metagenomic library. These proteins exhibit low to
moderate identity to known esterase sequences (<52%) and to each other (11.9 to 63.7%) and appear to have originated from
unknown species or from genera of Proteobacteria related to Thiothrix/Leucothrix (MGS-RG1/RG2) and to the Rhodobacteraceae
group (MGS-RG3). A library of 131 esters and 31 additional esterase/lipase preparations was used to evaluate the activity
profiles of these enzymes. All 3 of these enzymes had greater esterase than lipase activity and exhibited specific activities with
ester substrates (<356Umg 1) in the range of similar enzymes. MGS-RG3 was inhibited by salts and pressure and had a low
optimal temperature (30°C), and its substrate profile clustered within a group of low-activity and substrate-restricted marine
enzymes. In contrast, MGS-RG1 and MGS-RG2 were most active at 45 to 50°C and were salt activated and barotolerant. They
also exhibited wider substrate profiles that were close to those of highly active promiscuous enzymes from a marine hydrothermal
vent (MGS-RG2) and from a cold brackish lake (MGS-RG1). The data presented are discussed in the context of promoting
the examination of enzyme activities of taxa found in habitats that have been neglected for enzyme prospecting; the enzymes
found in these taxa may reflect distinct habitat-specific adaptations and may constitute new sources of rare reaction specificities.The European
Community project MAMBA (FP7-KBBE-2008-226977), grant BIO2011-25012 from the Spanish Ministry of the
Economy and Competitiveness (formerly MICINN). P.N.G. and O.V.G.
were supported by EU FP7 project MICROB3 (FP7-OCEAN.2011
287589). This work received support from the Government of Canada
through Genome Canada and the Ontario Genomics Institute (grant
2009-OGI-ABC-1405 to A.F.Y. and A.S.) and from the U.S. National Institutes
of Health (grants GM074942 and GM094585 to A.S. through the
Midwest Center for Structural Genomics).http://aem.asm.orgam201
Systematic Genetic Screens Reveal the Dynamic Global Functional Organization of the Bacterial Translation Machinery
Bacterial protein synthesis is an essential, conserved, and environmentally responsive process. Yet, many of its components and dependencies remain unidentified. To address this gap, we used quantitative synthetic genetic arrays to map functional relationships among >48,000 gene pairs in Escherichia coli under four culture conditions differing in temperature and nutrient availability. The resulting data provide global functional insights into the roles and associations of genes, pathways, and processes important for efficient translation, growth, and environmental adaptation. We predict and independently verify the requirement of unannotated genes for normal translation, including a previously unappreciated role of YhbY in 30S biogenesis. Dynamic changes in the patterns of genetic dependencies across the four growth conditions and data projections onto other species reveal overarching functional and evolutionary pressures impacting the translation system and bacterial fitness, underscoring the utility of systematic screens for investigating protein synthesis, adaptation, and evolution
Decoding the ocean's microbiological secrets for marine enzyme biodiscovery
A global census of marine microbial life has been underway over the past several decades. During this period, there have been scientific breakthroughs in estimating microbial diversity and understanding microbial functioning and ecology. It is estimated that the ocean, covering 71% of the earth's surface with its estimated volume of about 2 x 10(18) m(3) and an average depth of 3800 m, hosts the largest population of microbes on Earth. More than 2 million eukaryotic and prokaryotic species are thought to thrive both in the ocean and on its surface. Prokaryotic cell abundances can reach densities of up to 10(12) cells per millilitre, exceeding eukaryotic densities of around 10(6) cells per millilitre of seawater. Besides their large numbers and abundance, marine microbial assemblages and their organic catalysts (enzymes) have a largely underestimated value for their use in the development of industrial products and processes. In this perspective article, we identified critical gaps in knowledge and technology to fast-track this development. We provided a general overview of the presumptive microbial assemblages in oceans, and an estimation of what is known and the enzymes that have been currently retrieved. We also discussed recent advances made in this area by the collaborative European Horizon 2020 project 'INMARE'
Pressure adaptation is linked to thermal adaptation in salt-saturated marine habitats
The present study provides a deeper view of protein
functionality as a function of temperature, salt and
pressure in deep-sea habitats. A set of eight different
enzymes from five distinct deep-sea (3040–4908 m
depth), moderately warm (14.0–16.5°C) biotopes,
characterized by a wide range of salinities (39–348
practical salinity units), were investigated for this
purpose. An enzyme from a ‘superficial’ marine
hydrothermal habitat (65°C) was isolated and characterized
for comparative purposes. We report here the
first experimental evidence suggesting that in saltsaturated
deep-sea habitats, the adaptation to high
pressure is linked to high thermal resistance (P
value = 0.0036). Salinity might therefore increase the
temperature window for enzyme activity, and possibly
microbial growth, in deep-sea habitats. As an
example, Lake Medee, the largest hypersaline deepsea
anoxic lake of the Eastern Mediterranean Sea,
where the water temperature is never higher than
16°C, was shown to contain halopiezophilic-like
enzymes that are most active at 70°C and with denaturing
temperatures of 71.4°C. The determination of
the crystal structures of five proteins revealed
unknown molecular mechanisms involved in protein
adaptation to poly-extremes as well as distinct active
site architectures and substrate preferences relative
to other structurally characterized enzymes.European Community project MAMBA (FP7-KBBE-2008-226977). This grant BIO2011-25012 from the Spanish Ministry of Economy and Competitiveness (formerly MICINN). European Commission for ‘MicroB3’ grant (FP7-OCEAN.2011-2 (contract Nr
287589)). Government of Canada through Genome Canada
and the Ontario Genomics Institute (grant 2009-OGI-ABC-1405) and
U.S. National Institutes of Health (grants GM074942 and GM094585). Midwest Center for Structural Genomics).http://onlinelibrary.wiley.com/journal/10.1111/(ISSN)1462-2920hb2016Biochemistr