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Laccases are enzymes from the multi-copper oxidase family that have gathered a lot of attention due to their wide range of substrates, their interspecies variability and their still elusive mechanism of action. The presence of four copper atoms in their active site makes them an interesting model for the study of the relationship between the structure and function of proteins. It is possible to find them in fungi, plants and prokaryotes. Bacterial laccases display many advantages over fungal laccases in terms of their application. They have, in general, a greater thermal stability and a different pH profile, which contributes to widening their field of possible application. In the present work, a novel laccase from an Antarctic microorganism, Geobacillus sp. ID17, is purified and characterized. This is the first Antarctic bacterial laccase to be functionally described. It was found to be active at neutral pH and to have greater activity at 55°C. Its catalytic constants are in the order of other bacterial laccases. Screening for different potential substrates was also performed, showing that this novel laccase is more selective than commercial laccases. This enzyme could find potential application in the generation of gallic acid polymers or in organic synthesis in contexts where meticulous substrate discrimination is needed.

Microorganisms can play a significant role in material corrosion, with bacterial biofilms as major participants in microbially influenced corrosion (MIC). The exact mechanisms by which this takes place are poorly understood, resulting in a scarcity of information regarding MIC detection and prevention. In this work, a consortium of moderately thermophilic bacteria isolated from a biofilm growing over aluminum alloy 7075 was characterized. Its effect over the alloy was evaluated on a 40-day period using Electron Microscopy, demonstrating acceleration of corrosion in comparison to the abiotic control. The bacterial consortium was biochemically and microbiologically characterized as an as an attempt to elucidate factors contributing to corrosion. Molecular analysis revealed that the consortium consisted mainly of members of the Bacillusgenus, with lower abundance of other genera such as Thermoanaerobacterium, Anoxybacillus and Paenibacillus. The EPS polysaccharide presented mainly mannose, galactose, rhamnose and ribose. Our observations suggest that the acidification of the culture media resulting from bacterial metabolism acted as the main contributor to corrosion, hinting at an unspecific mechanism. The consortium was not sulfate-reducing, but it was found to produce hydrogen, which could also be a compounding factor for corrosion.

Laccases are enzymes able to catalyze the oxidation of a wide array of phenolic and non-phenolic compounds using oxygen as co-substrate and releasing water as by-product. They are well known to have wide substrate specificity and in recent years, have gained great biotechnological importance. To date, most well studied laccases are from fungal and mesophilic origin, however, enzymes from extremophiles possess an even greater potential to withstand the extreme conditions present in many industrial processes. This research work presents the heterologous production and characterization of a novel laccase from a thermoalkaliphilic bacterium isolated from a hot spring in a geothermal site. This recombinant enzyme exhibits remarkably high specific activity (>450,000 U/mg) at 70 °C, pH 6.0, using syringaldazine substrate, it is active in a wide range of temperature (20–90 °C) and maintains over 60% of its activity after 2 h at 60 °C. Furthermore, this novel spore-coat laccase is able to biodecolorize eight structurally different recalcitrant synthetic dyes (Congo red, methyl orange, methyl red, Coomassie brilliant blue R250, bromophenol blue, malachite green, crystal violet and Remazol brilliant blue R), in just 30 min at 40 °C in the presence of the natural redox mediator acetosyringone.

Two extracellular xylanases, denominated X2 and X3, were purified and characterized from the halotolerant bacterium Bacillus sp. Asc6BA isolated from “Salar de Ascotan” in the Atacama Desert. Xylanases were purified by anion exchange, cation exchange and size exclusion liquid chromatography. Xylanase X2 and X3 were purified ~ 690-fold and ~ 629-fold, respectively, compared to the concentrated extracellular fraction with a final specific activity of 169 and 154 u mg−1, respectively. Optimal conditions of pH and temperature of xylanolytic activity were 6.0 and 60 °C for X2 and 7.0 and 60 °C for X3. Half-life of X2 xylanase was 30 min at 50 °C, while X3 xylanase was remarkably more thermostable, retaining more than 70% of its activity after 32 h of incubation at 50 °C. X2 exhibited Km, Vmax and kcat values of 7.17 mg mL−1, 1.28 mM min−1 mg−1 and 425.33 s−1, respectively. X3 exhibited Km, Vmax and kcat values of 6.00 mg mL−1, 19.25 mM min−1 mg−1 and 82,515 s−1, respectively. In addition to their thermal stabilities, these enzymes were shown to be resistant to freeze-drying. These stability properties, in addition to the ability of these enzymes to be active in a wide range of temperatures and pHs, make these xylanases good candidates for industrial applications.

Violacein is an intensely purple pigment synthesized by various genera of bacteria that has been discovered to have a wide range of interesting biological activities which range from anticarcinogenic to antibacterial. One of the hindrances for its real-life application is that the first microorganisms found to produce the compound may act as opportunistic pathogens. Here, we report the isolation and characterization of violacein from a non-pathogenic Antarctic Iodobacter strain. Its anti-microbial properties were also tested. The method proposed here for the purification of violacein shows high yields, indicating that this Antarctic microorganism could be a valuable source for this important pigment. This is the first characterization of violacein from an Antarctic Iodobacter strain and here we also present a viable method to obtain this pigment for potential biotechnological applications.

The present work describes the results of the preliminary characterization of a novel enzyme with catalase activity isolated from psychrotolerant bacteria. Catalases decompose hydrogen peroxide (H2O2) to water and oxygen. Chemical immobilization is the best solution in case of reactions in which a gaseous product is created hence immobilization via divinyl sulfone having used a polymeric regenerated cellulose membrane was proposed. The preparations were checked according to their activity and stability. In the textile industry, a catalase is used to decompose the hydrogen peroxide waste after bleaching fabrics. During photocatalytic oxidation, H2O2 is added at relatively high concentration app. 20–80 g/L and the pH of these solutions is in the range between 7 and 10. The psychrophilic catalase described in this work presents high activity up to 40 g/L at pH 7–10, while a commercial alternative isolated from bovine liver only displays activity up to 14 g/L at pH not higher than 9. The stability of enzymatic activity is strongly dependent on temperature and is also a function of substrate concentration. The results obtained with this psychrophilic catalase are promising for textile effluence treatment.

New experimental techniques and protocols are needed to study microbiologically influenced corrosion (MIC). In this work, we studied the capacity of microorganisms to modify their environment in vitro. The methodology involved cyclic voltammetry measurements using carbon electrodes modified with cobalt phthalocyanine, which is a known catalyst for the two-electron reduction of O2 to give peroxide. Mueller-Hinton (21 g/L) was used as an electrolyte in a sterilized aqueous medium. The open circuit potential (OCP) and oxygen concentration behaved similarly over time, with generally similar growth curves. However, there were peculiarities that indicated the presence of peroxide and catalase. Catalase activity was demonstrated by comparing the voltammetric responses of the culture medium in the absence and presence of bacterial strains of Escherichia coli (Gram +) and Staphylococcus aureus (Gram-), both facultative and catalase positive. With this system, which is capable of discriminating O2 and hydrogen peroxide, catalase activity is highly evident, and at its maximum at the end of the exponential stage of the growth curve.

Ecological and efficient alternatives to industrial processes have sparked interest for using microorganisms and enzymes as biocatalysts. One of the difficulties is finding candidates capable of resisting the harsh conditions in which industrial processes usually take place. Extremophiles, microorganisms naturally found in "extreme" ecological niches, produce robust enzymes for bioprocesses and product development. Thermophiles like Geobacillus, Alyciclobacillus, Anoxybacillus, Pyrococcus and Thermoccocus are some of the extremophiles containing enzymes showing special promise for biocatalysis. Glutamate dehydrogenase used in food processes, laccases and xylanases in pulp and paper processes, nitrilases and transaminases for pharmaceutical drug synthesis and lipases present in detergents, are examples of the increasing use of enzymes for biocatalytic synthesis from thermophilic microorganisms. Some of these enzymes from thermophiles have been expressed as recombinant enzymes and are already in the market. Here we will review recent discoveries of thermophilic enzymes and their current and potential applications in industry.

Silver nanoparticles (AgNPs) have been widely recognized as antibacterial agents. However, its stability and activity over time have been poorly studied. In this work, the properties and characteristics of differently stabilized AgNPs were evaluated during a span of time. The surface capping agents were diclofenac (d), and ketorolac (k), which currently are used as anti-inflammatory in human medicine. On evaluating the size variation over time, it was observed that the AgNPs-k are the most stable, unlike the non-capped nanoparticles agglomerate and precipitate. UV–Vis spectroscopy analysis showed that the absorbance during time decreases for the three types of nanoparticles, but the decrease is less marked for the two types of anti-inflammatory-capped AgNPs. The rapid loss of the optical prop- erties of bare AgNPs, is mainly due to oxidation, agglomeration, and precipitation of this nanoparticles. The potential cytotoxicity of the AgNPs, evaluated through the formation of the superoxide anion using XXT, showed that both, AgNPs-k and AgNPs-d, generate the radical anion when the samples are irradiated with UV light at 365 nm. This effect appears associated with the capping agents, since the bare nanoparticles did not promote the formation of the superoxide anion. The antibacterial activity of the AgNPs throughout time, against two microorganisms (Escherichia coli and Staphylococcus aureus), was also evaluated. The results showed that capping agents played a decisive role in the antibacterial ability of AgNPs and also in enhancing the antibacterial activity over time.

The Orca Seamount is a submarine volcanic structure, located in a tectonic area of the Bransfield Strait, characterized by cortical extension and roll back-type subduction. Recent investigations have described the presence of hydrothermal activity and thermophilic microorganisms in this submarine volcano, raising questions regarding the role these microorganisms might play in the environment. The presence of hydrothermal activity interacting with cold Antarctic marine waters has probably exerted a great impact on the chemistry of the Orca Seamount area, providing different types of substrates capable to support complex microbial communities. In this work, we further study the Orca Seamount area with respect to the mineralogy present in this environment and the role microorganisms might play in the biogeochemical cycles. Here we show that the assemblage of minerals detected in the Orca Seamount area is like those commonly found in other hydrothermal environments, consistent with previous investigations reporting hydrothermal activity in this zone. Sulfur- and iron-bearing minerals in addition to inorganic soluble compounds are able to support chemosynthetic microbial communities inhabiting the Orca Seamount. The role of these microorganisms on the sulfur, iron, and carbon cycle is discussed and analyzed in the context of the mineralogy and conditions of the environment.

A novel hyperthermophilic crenarchaeon, strain 3507LTT, was isolated from a terrestrial hot spring near Tinguiririca volcano, Chile. Cells were non-motile thin, slightly curved filamentous rods. It grew at 73–93 °C and pH range of 5 to 7.5 with an optimum at 85 °C and pH 6.0–6.7. The presence of culture broth filtrate of another hyperthemophilic archaeon as well as yeast extract was obligatory for growth of the novel isolate. Strain 3507LTT is an anaerobic chemoorganoheterotroph, fermenting monosaccharides, disaccharides and polysaccharides (lichenan, starch, xanthan gum, xyloglucan, alpha-cellulose and amorphous cellulose). No growth stimulation was detected when nitrate, thiosulfate, selenate or elemental sulfur were added as the electron acceptors. The complete genome of strain 3507LTT consisted of a single circular chromosome with size of 1.63 Mbp. The DNA G+C content was 53.9%. According to the 16S rRNA gene sequence as well as conserved protein sequences phylogenetic analyses, strain 3507LTT together with Thermofilum uzonense formed a separate cluster within a Thermofilaceae family (Thermoproteales/Thermoprotei/Crenarchaeota). Based on phenotypic characteristics, phylogeny as well as AAI comparisons, a novel genus and species Infirmifilum lucidum strain 3507LTT (=VKM B-3376T = KCTC 15797T) gen. nov. sp. nov. was proposed. Its closest relative, Thermofilum uzonense strain 1807–2T should be reclassified as Infirmifilum uzonense strain 1807–2T comb. nov. Finally, based on phylogenomic and comparative genome analyses of representatives of Thermofilaceae family and other representatives of Thermoproteales order, a proposal of transfer of the family Thermofilaceae into a separate order Thermofilales ord. nov. was made.

Two extracellular xylanases, denominated X2 and X3, were purified and characterized from the halotolerant bacterium Bacillus sp. Asc6BA isolated from "Salar de Ascotán" in the Atacama Desert. Xylanases were purified by anion exchange, cation exchange and size exclusion liquid chromatography. Xylanase X2 and X3 were purified ~ 690-fold and ~ 629-fold, respectively, compared to the concentrated extracellular fraction with a final specific activity of 169 and 154 u mg

Background: Xylan is the second most abundant polysaccharide biomass on the earth, the polymers have a backbone of β-1,4-linked xylose residues with various side-chain substitutions, such as arabinose, acetic acid, glucuronic acid, and other esterified groups, thus, the removal of arabinose side groups by α-L-arabinofuranosidases is helpful in various industrial processes involving xylan treatment. Bacillus subtilis ATCC 6051a is known for its excellent capacity of secretory production of recombinant peptides, however, poor experience in genetic manipulation and lack of universal expression elements impede this strain for wider application.Results: Xylose inducible comK was integrated into the genome of B. subtilis ATCC 6051a, and the transformation efficiency of the engineered strain B. subtilis 164S was increased by more than 1000 folds. B. subtilis 164S was further modified to generate B. subtilis 164T7P which incorporates a D-xylose inducible T7 RNA polymerase. The recombinant GFP expressed by 164T7P is more than thirteen times that achieved by P43 promoter, representing the most efficient expression system that had been ever reported in B. subtilis . Subsequently, abfA1 , encoding a glycoside hydrolase (GH) family 51 enzyme was cloned and overexpressed in 164T7P. The activity of recombinant α-L-arabinofuranosidase (AbfA1) reached 90.6±2.0 U mL -1 in the fermentation broth. Using p NPA as a substrate, kinetic parameters of the crude enzyme were Km of 1.4±0.1 mM and kcat of 139.4 s −1 . The optimum temperature and pH of the recombinant AbfA1 towards p NPA were observed to be 45 °C and pH 6.5, respectively.Conclusion: With enhanced cellular competence and introduction of T7 RNA polymerase, B. subtilis ATCC 6051a was engineered as a versatile cell tool for recombinant production of heterologous peptides employing T7 promoter. The novel expression kit demonstrated a very low level of leaky expression of target genes. The efficiency and applicability of the system were demonstrated by high-level production of a bacterial type α-L-arabinofuranosidase.

Inhibiting pathogenic bacterial adherence on surfaces is an ongoing challenge to prevent the development of biofilms. Multilayer polyelectrolyte films are feasible antibacterial materials. Here, we have designed new films made of carbohydrate polyelectrolytes to obtain antibacterial coatings that prevent biofilm formation. The polyelectrolyte films were constructed from poly(maleic anhydride- alt-styrene) functionalized with glucofuranose derivatives and quaternized poly(4-vinylpyridine) N-alkyl. These films prevent Pseudomonas aeruginosa and Salmonella Typhimurium, two important bacterial contaminants in clinical environments, from adhering to surfaces. When the film was composed of more than 10 layers, the bacterial population was greatly reduced, while the bacteria remaining on the film were morphologically damaged, as atomic force microscopy revealed. The antibacterial capacity of the polyelectrolyte films was determined by the combination of thickness, wettability, surface energy, and most importantly, the conformation that polyelectrolytes adopt the function of nature of the carbohydrate group. This polyelectrolyte film constitutes the first green approach to preventing pathogenic bacterial surface adherence and proliferation without killing the bacterial pathogen.

A recombinant laccase from thermophilic bacteria was used to construct an enzyme–multi-walled carbon nanotube composite catalyst, which shows improved stability as a bio-electrocatalyst for the oxygen reduction reaction compared to similar conjugates prepared using the commercially available T. versicolor laccase.

This article reports the electrochemical studies of bio-electrocatalytic oxygen reduction reaction catalyzed by a new laccase from a thermophilic bacterium, which exhibits superior stability and activity retention compared to fungal laccases. The electro-kinetic study of this catalytic reaction was conducted using a classic tool of electrochemistry, the rotating ring disk electrode (RRDE). Bacillus sp. FNT laccase was immobilized onto a multiwall carbon nanotube modified glassy carbon disk electrode using a molecular tethering agent, 1-pyrenebutanoic acid succinimidyl ester (PBSE). The conditions for laccase immobilization were optimized to prepare a highly active bioconjugate for the electrochemical reduction of oxygen. The kinetic parameters such as Tafel slopes, number of electrons transferred, electrochemical rate constant and the electron transfer rate were calculated from RRDE measurements. The rate constant of the four-electron transfer reaction was calculated to be 3.21 x 10(-1) +/- 8.5 x 10(-3) cm s(-1). The slope value obtained from Tafel plot was close to that of value for the ideal four-electron transfer oxygen reduction, suggesting excellent electro-catalytic activity of the FNT laccase-MWCNT conjugates. (C) 2018 The Electrochemical Society.

This work studied the microbiologically influenced corrosion susceptibility of the AA2024-T3 alloy exposed to sterile and inoculated media with three coccoidal bacteria, which were isolated from the corrosion product of a fuel outlet, belonging to a Twin Otter aircraft exposed to Antarctic environmental conditions. The consortium was mainly constituted by three bacteria, which have a close affiliation to Kocuria rhizophila, Staphylococcus epidermidis and Staphylococcus aureus. Electrochemical methods and surface analysis were carried out to investigate the corrosion susceptibility of the alloy under study in the presence of these bacteria. Electrochemical analysis suggested that the protective character of the oxide of the alloy film was modified during the immersion time in all media. Moreover, the surface analysis showed also that these bacteria influenced homogeneity of this oxide. In particular, the K. rhizophila increased slightly the current and decreased the resistance of AA2024-T3 against corrosion, instead the S. epidermidis showed a slight protective effect, while non-significant effect was observed with the presence of the consortium in the medium.

Background Antifreeze proteins (AFPs) production is a survival strategy of psychrophiles in ice. These proteins have potential in frozen food industry avoiding the damage in the structure of animal or vegetal foods. Moreover, there is not much information regarding the interaction of Antarctic bacterial AFPs with ice, and new determinations are needed to understand the behaviour of these proteins at the water/ice interface. Results Different Antarctic places were screened for antifreeze activity and microorganisms were selected for the presence of thermal hysteresis in their crude extracts. Isolates GU1.7.1, GU3.1.1, and AFP5.1 showed higher thermal hysteresis and were characterized using a polyphasic approach. Studies using cucumber and zucchini samples showed cellular protection when samples were treated with partially purified AFPs or a commercial AFP as was determined using toluidine blue O and neutral red staining. Additionally, genome analysis of these isolates revealed the presence of genes that encode for putative AFPs. Deduced amino acids sequences from GU3.1.1 (gu3A and gu3B) and AFP5.1 (afp5A) showed high similarity to reported AFPs which crystal structures are solved, allowing then generating homology models. Modelled proteins showed a triangular prism form similar to β-helix AFPs with a linear distribution of threonine residues at one side of the prism that could correspond to the putative ice binding side. The statistically best models were used to build a protein-water system. Molecular dynamics simulations were then performed to compare the antifreezing behaviour of these AFPs at the ice/water interface. Docking and molecular dynamics simulations revealed that gu3B could have the most efficient antifreezing behavior, but gu3A could have a higher affinity for ice. Conclusions AFPs from Antarctic microorganisms GU1.7.1, GU3.1.1 and AFP5.1 protect cellular structures of frozen food showing a potential for frozen food industry. Modeled proteins possess a β-helix structure, and molecular docking analysis revealed the AFP gu3B could be the most efficient AFPs in order to avoid the formation of ice crystals, even when gu3A has a higher affinity for ice. By determining the interaction of AFPs at the ice/water interface, it will be possible to understand the process of adaptation of psychrophilic bacteria to Antarctic ice. Electronic supplementary material The online version of this article (doi:10.1186/s12934-017-0737-2) contains supplementary material, which is available to authorized users.

Background: A moderately thermophilic, slightly halophilic, aerobic, Gram-stain negative, bacterial strain, SLM16, was isolated from a mixed of seawater–sand-sediment sample collected from a coastal fumarole located in Whalers Bay, Deception Island, Antarctica. The aim was to screen for thermophilic microorganisms able to degrade primary amines and search for amine transaminase activity for potential industrial application. Results: Identification and partial characterization of the microorganism SLM16 were carried out by means of morphological, physiological and biochemical tests along with molecular methods. Cells of strain SLM16 were non-motile irregular rods of 1.5–2.5 μm long and 0.3–0.45 μm wide. Growth occurred in the presence of 0.5–5.5% NaCl within temperature range of 35–55 °C and pH range of 5.5–9.5, respectively. The DNA G+C composition, estimated from ftsY gene, was 66% mol. Phylogenetic analysis using de 16S rRNA gene sequence showed that strain SLM16 belongs to the marine bacterial genus Albidovulum. Conclusion: Strain SLM16 is a moderate thermophilic Gram negative microorganisms which belongs to the marine bacterial genus Albidovulum and is closely related to Albidovulum inexpectatum species based on phylogenetic analysis. Additionally, amine-transaminase activity towards the arylaliphatic amine α-methylbenzylamine was detected.

Common alloys used for the manufacture of aircrafts are subject to different forms of environmental deterioration. A major one is corrosion, and there is a strong body of evidence suggesting that environmental microorganisms initiate and accelerate it. The development of an appropriate strategy to reduce this process depends on the knowledge concerning the factors involved in corrosion. In this work, a biofilm forming bacterial consortium was extracted in situ from the corrosion products formed in an aircraft exposed to Antarctic media. Two thermophilic bacteria, an Anoxybacillus and a Staphylococcus strain, were successfully isolated from this consortium. Two extracellular enzymes previously speculated to participate in corrosion, catalase and peroxidase, were detected in the extracellular fraction of the consortium. Additionally, we assessed the individual contribution of those thermophilic microorganisms on the corrosion process of 7075-T6 aluminum alloy, which is widely used in aeronautical industry, through electrochemical methods and surface analysis techniques.

Microorganisms present in Antarctica have to deal not only with cold temperatures but also with other environmental conditions, such as high UV radiation, that trigger the generation of reactive oxygen species. Therefore, Antarctic microorganisms must have an important antioxidant defense system to prevent oxidative damage. One of these defenses are antioxidant enzymes, such as catalase, which is involved in the detoxification of hydrogen peroxide produced under oxidative conditions. Here, we reported the isolation and partial characterization of an Antarctic bacterium belonging to the Serratia genus that was resistant to UV-C radiation and well-adapted to cold temperatures. This microorganism, denominated strain I1P, was efficient at decreasing reactive oxygen species levels produced after UV-C irradiation. Genomic and activity assays suggested that the enzymatic antioxidant defense mechanisms of strain I1P, especially its catalase enzyme, may confer UV resistance. This catalase was active in a wide range of temperatures (20&ndash, 70 &deg, C), showing optimal activity at 50 &deg, C (at pH 7.0), a remarkable finding considering its psychrotolerant origin. In addition, this enzyme was thermostable, retaining around 60% of its activity after 6 h of incubation at 50 &deg, C. The antioxidant defense systems of strain I1P, including its surprisingly thermoactive and thermostable catalase enzyme, make this microorganism a good source of biocompounds with potential biotechnological applications.

Background: The Antarctic continent is a source of extreme microorganisms. Millions of years of isolation have produced unique biodiversity with adaptive responses to its extreme environment. Although the Antarctic climate is mainly cold, the presence of several geothermal sites, including thermal springs, fumaroles, hot soils and hydrothermal vents, provides ideal environments for the development of thermophilic and hyperthermophilic microorganisms. Their enzymes, called thermoenzymes, are the focus of interest in both academic and industrial research, mainly due to their high thermal activity and stability. Glutamate dehydrogenase, is an enzyme that plays a key role in the metabolism of carbon and nitrogen catalyzing reversibly the oxidative deamination of glutamate to alpha-ketoglutarate and ammonium. It belongs to the family of oxidoreductases, is widely distributed and it has been highly regarded for use as biosensors, particularly for their specificity and ability to operate in photochemical and electrochemical systems. However, the use of enzymes as biosensors is relatively problematic due to their instability to high temperatures, organic solvents and denaturing agents. The purpose of this study is to present the partial characterization of a thermophilic microorganism isolated from Deception Island, Antarctica, that displays glutamate dehydrogenase activity. Results: In this work, we report the isolation of a thermophilic microorganism called PID15 from samples of Deception Island collected during the Antarctic Scientific Expedition ECA 46. This microorganism is a thermophile that grows optimally at 50 °C and pH 8.0. Scanning electron microscopy shows rod cells of 2.0 to 8.0 μm of length. Phylogenetic analysis of 16S rRNA gene revealed that this microorganism is closely related to Bacillus gelatini. This microorganism contains a thermostable glutamate dehydrogenase with optimal activity at pH 8.0 and temperatures for its activity from 37 to 50 °C, range of temperature of interest for biotechnological applications. This glutamate dehydrogenase is a highly thermostable enzyme. Conclusion: This is the first report of a microorganism from Antarctica containing a thermostable glutamate dehydrogenase that maintains its activity in a broad range of temperatures making it of potential interest for biotechnological applications.

The existence of multiple-antibiotic-resistant strains of environmental bacteria is commonly linked to human activities. However, multiple-antibiotic-resistant strains of bacteria are also widely found in the Antarctic that has limited human activity. This study was conducted to examine the prevalence of antibiotic-resistant strains among Antarctic bacteria. Forty-five bacterial strains from Estrellas lake of King George Island and Crater lake of Deception Island, Antarctic, were exposed to 30 different antibiotics. Forty out of the 45 bacterial strains were affiliated to 12 genera, Aeromicrobium, Arthrobacter, Bacillus, Brevundimonas, Cryobacterium, Dyadobacter, Flavobacterium, Methylibium, Pedobacter, Pseudomonas, Rhodococcus, and Sphingomonas. Among the bacteria, 43 strains were resistant to at least three antibiotics, and 26 strains were resistant to 10 or more different antibiotics. Pseudomonas spp. and four unknown Microbacteriaceae bacteria were found to be resistant to majority of the antibiotics tested. Two bacteria, each from Estrellas and Crater lakes, were sensitive to all the antibiotics tested. These results indicated that Antarctic bacteria are probably the reservoirs for antibiotic resistance genes.

In this paper we studied the influence of bacterial consortia on corrosion of fuel tanks of aircraft used often for pilot training. In the first part, we proceeded to the isolation and characterization of the consortia by DNA sequencing. The consortium was mainly constituted by two bacteria, which have a close affiliation to Bacillus subtilis and to Bacillus circulans . The substrate used in fuel tanks was an aluminum alloy 2024. Electrochemical measurements and morphological characterization were performed for different immersion times (1, 2, 3, 7 and 14 days) in sterile minimal salt medium (MSM) and in inoculated media (MSM + B. subtilis and MSM + consortia). For all immersion times, impedance diagrams showed a capacitive behavior characterized by a single constant time. Although for short exposure times, no corrosion process was evidenced, for longer exposure time, the resistance of the aluminum oxide ( R ox ) decreased markedly, indicating that a corrosion process was taking place.

To date, many industrial processes are performed using chemical compounds, which are harmful to nature. An alternative to overcome this problem is biocatalysis, which uses whole cells or enzymes to carry out chemical reactions in an environmentally friendly manner. Enzymes can be used as biocatalyst in food and feed, pharmaceutical, textile, detergent and beverage industries, among others. Since industrial processes require harsh reaction conditions to be performed, these enzymes must possess several characteristics that make them suitable for this purpose. Currently the best option is to use enzymes from extremophilic microorganisms, particularly archaea because of their special characteristics, such as stability to elevated temperatures, extremes of pH, organic solvents, and high ionic strength. Extremozymes, are being used in biotechnological industry and improved through modern technologies, such as protein engineering for best performance. Despite the wide distribution of archaea, exist only few reports about these microorganisms isolated from Antarctica and very little is known about thermophilic or hyperthermophilic archaeal enzymes particularly from Antarctica. This review summarizes current knowledge of archaeal enzymes with biotechnological applications, including two extremozymes from Antarctic archaea with potential industrial use, which are being studied in our laboratory. Both enzymes have been discovered through conventional screening and genome sequencing, respectively.

To obtain new insights into community compositions of hyperthermophilic microorganisms, defined as having optimal growth temperatures of 80 degrees C and above, sediment and water samples were taken from two shallow marine hydrothermal vents (I and II) with temperatures of 100 degrees C at Vulcano Island, Italy. A combinatorial approach of denaturant gradient gel electrophoresis (DGGE) and metagenomic sequencing was used for microbial community analyses of the samples. In addition, enrichment cultures, growing anaerobically on selected polysaccharides such as starch and cellulose, were also analyzed by the combinatorial approach. Our results showed a high abundance of hyperthermophilic archaea, especially in sample II, and a comparable diverse archaeal community composition in both samples. In particular, the strains of the hyperthermophilic anaerobic genera Staphylothermus and Thermococcus, and strains of the aerobic hyperthermophilic genus Aeropyrum, were abundant. Regarding the bacterial community, e-Proteobacteria, especially the genera Sulfurimonas and Sulfurovum, were highly abundant. The microbial diversity of the enrichment cultures changed significantly by showing a high dominance of archaea, particularly the genera Thermococcus and Palaeococcus, depending on the carbon source and the selected temperature.

Nitriles are important chemical building blocks for the synthesis of intermediates in fine chemical and pharmaceutical industries. Here, we report a new highly thermostable nitrilase from an Antarctic Pyrococcus sp. MC-FB, a hyperthermophilic archaeon. A gene that encoded a nitrilase was identified and subsequently cloned and overexpressed in Escherichia coli. The recombinant nitrilase, named NitMC-FB, is active as a homodimer (60 kDa) with an optimal temperature and pH of 90 °C and 7.0, respectively. NitMC-FB hydrolyzes preferentially aromatic nitriles, being the first aromatic nitrilase from an archaeon described so far. The K M and V max parameters were determined to be 13.9 mM and 3.7 μmol/min*mg, respectively, with 2-cyanopyridine as the substrate. Additionally, the recombinant nitrilase is highly thermostable with a half-life of 8 h at 90 °C.

The production of ever more complex fine chemicals for life sciences (health care, agrochemicals, nutraceuticals) requires selective synthetic biotechnological tools, which complement purely organic chemical synthesis with a rewarding economic and ecological impact. Biotechnology has become an important contributor to the gross domestic product in many countries, and this contribution translates also into enzyme applications for different markets and products, novel biotech companies, and international research programs and consortia. The present chapter deals with the different aspects of enzyme biocatalysis for chemical transformations, starting with the prospection and identification of novel microbial enzymes, which are becoming indispensable for current industrial processes. The benefits of extremophilic bioprospection and protein engineering along with the preponderant role of nature itself as the best source for finding truly novel enzymes with potential industrial applicability are discussed. Also, the different production processes, using immobilized and free biocatalysts for fine and bulk chemical production, are analyzed. Finally, new reactions and developments in enzymatic catalysis will be presented as well as green chemistry aspects as an inherent feature of industrial biotransformation, where novel technologies for cofactor regeneration and solvents for biotransformations are featured as “green” solutions.

Amine-transaminases (ATAs) are enzymes that catalyze the reversible transfer of an amino group between primary amines and carbonyl compounds. They have been widely studied in the last decades for their application in stereoselective synthesis of chiral amines, which are one of the most valuable building blocks in pharmaceuticals manufacturing. Their excellent enantioselectivity, use of low-cost substrates and no need for external cofactors has turned these enzymes into a promising alternative to the chemical synthesis of chiral amines. Nevertheless, its application at industrial scale remains limited mainly because most of the available ATAs are scarcely tolerant to harsh reaction conditions such as high temperatures and presence of organic solvents. In this work, a novel (S)-ATA was discovered in a thermophilic bacterium, Albidovulum sp. SLM16, isolated from a geothermal Antarctic environmental sample, more specifically from a shoreline fumarole in Deception Island. The transaminase-coding gene was identified in the genome of the microorganism, cloned and overexpressed in Escherichia coli for biochemical characterization. The activity of the recombinant ATA was optimal at 65 °C and pH 9.5. Molecular mass estimates suggest a 75 kDa homodimeric structure. The enzyme turned out to be highly thermostable, maintaining 80% of its specific activity after 5 days of incubation at 50 °C. These results indicate that ATA_SLM16 is an excellent candidate for potential applications in biocatalytic synthesis. To the best of our knowledge, this would be the first report of the characterization of a thermostable (S)-ATA discovered by means of in vivo screening of thermophilic microorganisms.

Enzymes produced by extremophiles offer an attractive solution to industrial biocatalysis since unlike most industrial enzymes today, extremozymes are more stable and robust, therefore making them ideal candidates for industrial applications. For industry, confirming the functionality of a biocatalyst under the conditions of a particular process makes the difference between obtaining or not the required product. Although the molecular approaches allow access to genes of interest in a fast way, to obtain large amounts of functional proteins, it is not possible by the sole use of genomics. Additionally, these approaches cannot discern properties such as: activity, pH and temperature stability, and inhibitors, only possible to measure through a functional approach. New technologies will need to integrate the functional approach into the large-scale biotechnology by implementing the technological capacity to analyse some intrinsic features to search for various enzymatic activities at the same time. This chapter will analyse the most recent and relevant advances for the discovery and development of extremophilic enzymes with a special emphasis in the implementation of functional screening strategies as well as rapid and efficient discovery tools. Furthermore, a group of selected biotransformations will be analysed in the context of the applicability of extremozymes in an industrial context.

Antarctica is an extreme continent composed of cold environments but also of several geothermal sites, among them is Deception Island, an active stratovolcano located in the South Shetland archipelago. From this island, few microbiological studies have been performed, and the presence of archaea has not been reported. In order to investigate the archaeal diversity in hydrothermalism from Deception Island, different submarine samples were taken from the flooded caldera. Samples were analyzed by denaturing gradient gel electrophoresis (DGGE) of the 16S rRNA gene in conjunction with culture-dependent methods at hyperthermophilic temperatures. Analysis from DGGE band sequencing showed the presence of archaea belonging to the hyperthermophilic genus Thermococcus and different uncultured archaea closely related to environmental clones from hydrothermal vents. Archaea from the psychrotolerant genus Methanococcoides were also detected. Additionally, we have successfully isolated an anaerobic hyperthermophilic archaeon closely related to Thermococcuscelericrescens. Cells were irregular cocci with a diameter between 0.6 and 2 μm and grew at 50–90 °C and at a NaCl concentration of 1–5 %. Here, we present, based on culture-dependent and culture-independent approaches, the first report on archaea from marine hydrothermal sites of Antarctica.