Extremophiles as a source for novel enzymes

Microbial life does not seem to be limited to specific environments. During the past few decades it has become clear that microbial communities can be found in the most diverse conditions, including extremes of temperature, pressure, salinity and pH. These microorganisms, called extremophiles, produce biocatalysts that are functional under extreme conditions. Consequently, the unique properties of these biocatalysts have resulted in several novel applications of enzymes in industrial processes. At present, only a minor fraction of the microorganisms on Earth have been exploited. Novel developments in the cultivation and production of extremophiles, but also developments related to the cloning and expression of their genes in heterologous hosts, will increase the number of enzyme-driven transformations in chemical, food, pharmaceutical and other industrial applications.     

Extremophiles as a source of novel enzymes for industrial application

Extremophilic microorganisms are adapted to survive in ecological niches such as at high temperatures, extremes of pH, high salt concentrations and high pressure. These microorganisms produce unique biocatalysts that function under extreme conditions comparable to those prevailing in various industrial processes. Some of the enzymes from extremophiles have already been purified and their genes successfully cloned in mesophilic hosts. In this review we will briefly discuss the biotechnological significance of extreme thermophilic (optimal growth 70–80 °C) and hyperthermophilic (optimal growth 85–100 °C) archaea and bacteria. In particular, we will focus on selected extracellular-polymer-degrading enzymes, such as amylases, pullulanases, cyclodextrin glycosyltransferases, cellulases, xylanases, chitinases, proteinases and other enzymes such as esterases, glucose isomerases, alcohol dehydrogenases and DNA-modifying enzymes with potential use in food, chemical and pharmaceutical industries and in environmental biotechnology. Received: 14 August 1998 / Received revision: 17 February 1999 / Accepted: 19 February 1999     

Soil phosphorus fractions and adsorption as affected by organic and inorganic sources

The effect of organic and inorganic sources of phosphorus (P) on soil P fractions and P adsorption was studied in a field without plant growth on a Kandiudalf in western Kenya. A high-quality organic source, Tithonia diversifolia (Hemsley) A. Gray leaves, and a low-quality source, maize (Zea mays L.) stover, were applied alone or in combination with triple superphosphate (TSP). The P rate was kept constant at 15 kg P ha^-1. Soil extractable P (resin, bicarbonate and sodium hydroxide), microbial biomass P and C and P adsorption isotherms were determined during 16 weeks after application of treatments. Application of tithonia either alone or with TSP increased resin P, bicarbonate P, microbial P, and sodium hydroxide inorganic P. Tithonia alone reduced P adsorption at 2–16 weeks. Maize stover had no effect on any of the P fractions or P adsorption. At 8 weeks, the application of tithonia reduced microbial C-to-P ratio (20) as compared to maize stover, TSP and the control (31–34). The reduction in P adsorption by tithonia was accompanied by increases in all measured P fractions, the sum of P in those fractions (resin, bicarbonate and sodium hydroxide) being larger than the P added. The reduction in P adsorption apparently resulted from competition for adsorption sites, probably by organic anions produced during decomposition of the high quality tithonia. Integration of inorganic P (TSP) with organic materials had little added benefit compared to sole application of TSP, except that combination of tithonia with TSP increased microbial biomass. The results indicate that a high quality organic input can be comparable to or more effective than inorganic P in increasing P availability in the soil.     

Adaptivity of Archaeal and Bacterial Extremophiles

Microbiology - Extremophilic prokaryotes, inhabitants of hot, cold, acidic, alkaline, saline, and deep-sea ecosystems, are classified as mono- and polyextremophilic or extreme-tolerant. Under...     

Transport of Compatible Solutes in Extremophiles

Salt-tolerant as well as moderately halophilic and halophilic organisms have to maintain their turgor. One strategy is to accumulate small organic compounds, compatible solutes, by de novo synthesis or uptake. From a bioenergetic point of view, uptake is preferred over biosynthesis. The transport systems catalyzing uptake of compatible solutes are of primary or secondary nature and coupled to ATP hydrolysis or ion (H+, Na+) symport. Expression of the transporter genes as well as the activity of the transporters is regulated by salinity/osmolarity and one of the key questions is how salinity or osmolarity is sensed and the signal transmitted as far as to gene expression and transporter activation. Recent studies shed light on the nature and the activation mechanisms of solute transporters in extremophiles, and this review summarizes current knowledge on the structure, function and osmo- or salt-regulation of transporters for compatible solutes in extremophiles.     

Extremophiles as Plant Probiotics to Promote Germination and Alleviate Salt Stress in Soybean

Journal of Plant Growth Regulation - Bacteria isolated from extreme environments have been shown to promote plant growth under challenging conditions. This study aimed to examine the performance of...     

极端微生物：一种新型的酶资源

极端微生物具有自身独特的特点和代谢产物 ,在食品工业、化工、药用工业和环境生物技术领域都有潜在的应用。一些酶已经得到纯化 ,其基因在宿主中已成功克隆。主要介绍和讨论极端微生物的类型、基因组及极端酶类的生产、分离与应用。     

Pure culture growth of ectomycorrhizal fungi on inorganic nitrogen sources

Four ectomycorrhizal fungi were tested for their ability to grow (i.e., mycelial mat radial extension and fungal biomass) on nutrient media either supplemented with ammonium-nitrogen or nitrate-nitrogen or in the absence of an inorganic nitrogen source.Pisolithus tinctorius, Cenococcum geophilum andThelephora terrestris exhibited greater growth on ammonium-nitrogen.Suillus granulatus grew better on the nitrate-nitrogen nutrient medium. Regardless of inorganic nitrogen form preference (i.e., ammonium-nitrogen or nitrate-nitrogen), all 4 species showed some growth on each of the 3 nutrient media. Growth rate maxima varied by fungal species as well as by inorganic nitrogen source. Maximum growth rate forT. terrestris exceeded rates exhibited by the other 3 fungi by 2–5 times.     

Comparison of CO2 and bicarbonate as inorganic carbon sources for triacylglycerol and starch accumulation in Chlamydomonas reinhardtii

Microalgae are capable of accumulating high levels of lipids and starch as carbon storage compounds. Investigation into the metabolic activities involved in the synthesis of these compounds has escalated since these compounds can be used as precursors for food and fuel. Here, we detail the results of a comprehensive analysis of Chlamydomonas reinhardtii using high or low inorganic carbon concentrations and speciation between carbon dioxide and bicarbonate, and the effects these have on inducing lipid and starch accumulation during nitrogen depletion. High concentrations of CO₂ (5%; v/v) produced the highest amount of biofuel precursors, transesterified to fatty acid methyl esters, but exhibited rapid accumulation and degradation characteristics. Low CO₂ (0.04%; v/v) caused carbon limitation and minimized triacylglycerol (TAG) and starch accumulation. High bicarbonate caused a cessation of cell cycling and accumulation of both TAG and starch that was more stable than the other experimental conditions. Starch accumulated prior to TAG and then degraded as maximum TAG was reached. This suggests carbon reallocation from starch‐based to TAG‐based carbon storage. Biotechnol. Bioeng. 2013; 110: 87–96. © 2012 Wiley Periodicals, Inc.     

Homology Modeling and Comparative Profiling of Superoxide Dismutase Among Extremophiles: Exiguobacterium as a Model Organism

Superoxide dismutase (SOD), a well known antioxidant enzyme, is known to exert its presence across bacteria to humans. Apart from their well-known antioxidant defense mechanisms, their association with various extremophiles in response to various stress conditions is poorly understood. Here, we have discussed the conservation and the prevalence of SODs among 21 representative extremophiles. A systematic investigation of aligned amino acid sequences of SOD from all the selected extremophiles revealed a consensus motif D-[VLE]-[FW]-E-H-[AS]-Y-[YM]. To computationally predict the correlation of SOD with the various stress conditions encountered by these extremophiles, Exiguobacterium was selected as a model organism which is known to survive under various adverse extremophilic conditions. Interestingly, our phylogenetic study based on SOD homology revealed that Exiguobacterium sibiricum was one of the closest neighbors of Deinococcus radiodurans and Thermus thermophilus. Next, we sought to predict 3-D model structure of SOD for E. sibiricum (PMDB ID: 0078260), which showed >95 % similarity with D. radiodurans R1 SOD. The reliability of the predicted SOD model was checked by using various validation metrics, including Ramachandran plot, Z-score and normalized qualitative model energy analysis score. Further, various physicochemical properties of E. sibiricum SOD were calculated using different prominent resources.

Electronic supplementary material

The online version of this article (doi:10.1007/s12088-014-0482-8) contains supplementary material, which is available to authorized users.     

不同无机氮源对东海原甲藻生长的影响

在实验室条件下,研究了不同浓度、不同形态的氮(NaNO₃、NH₄Cl和NaNO₂)对东海原甲藻(Prorocentrum donghaiense)生长的影响。结果表明:在NH₄Cl浓度为5.20μmol·L^-1(N/P为8)时藻的比生长率最高,而N/P为32和100时,藻的生长明显受到抑制。在NaNO₃为氮源时,最适N/P为12(氮浓度为7.80μmol·L^-1)。而NaNO₂作氮源,N/P为16(10.40μmol·L^-1)时藻的比生长率最高,N/P为32和100时藻的生长也明显受到抑制。研究显示,东海原甲藻对无机氮NH₄Cl和NaNO₃和NaNO₂都可以利用,最适生长的N/P比范围在8～20之间,相对高的N/P(32、100)不利于东海原甲藻的生长。     

Versatile Solidified Nanofibrous Cellulose-Containing Media for Growth of Extremophiles

Solidified media that employ a porous matrix of nanofibrous cellulose are described. The physicochemical stability of the porous structure allows the development of solidified media that can support the growth of extremophiles, such as acidophilic Acidiphilium, alkaliphilic Bacillus, thermophilic Geobacillus and Thermus, alkalithermophilic Bacillus, and acidothermophilic Sulfolobus microbes. The cellulose-supported media have several advantages over agar- and gellan gum-derived media, including versatility and stability.Solidified media using agar as a solidifying agent are indispensable in microbiology. For solid cultures of mesophilic microorganisms, agar is an ideal solidifying agent and has been used essentially unchanged since it was first introduced in the late 19th century (2, 15). However, the situation is very different when it comes to culturing extremophiles on solidified media. For example, agar media are not suitable for culturing thermophiles and hyperthermophiles because the solidification of agar is thermoreversible at around 50 to 60°C (21), and the media are unstable at temperatures much above 70°C for extended periods (1). Culturing extremophiles on solidified media under acidic or alkaline conditions presents a similar problem of instability.We reported previously the use of porous plates made of nanofibrous cellulose for microbial culture (4). Detailed accounts of the preparation procedure, fine structure of the cellulose plate, and its application to culturing representative mesophilic microorganisms (Escherichia coli, Bacillus subtilis, and Saccharomyces cerevisiae) appeared in that paper.Interestingly, the porous structure of the cellulose plate has a structural robustness comparable to that of highly crystalline cellulose (6-8) despite its seemingly fragile structure (4). In situ optical microscopic observation in hot and compressed water (5, 18) revealed that the cellulose plate remained unchanged in water up to 260°C at 25 MPa (4, 7). This finding suggested that the cellulose plate could be used, in principle, as a versatile platform for developing solidified media that support the growth of extremophilic microbes under a wide range of extreme culture conditions. A proof-of-concept experiment was done by successfully culturing Thermus thermophilus on the cellulose plate at 80°C (4), but its true potential, especially its versatility, still remained to be corroborated.In this paper, we show that a wide variety of extremophiles, including an acidophile, an alkaliphile, thermophiles, an acidothermophile, and an alkalithermophile, can be cultured on the cellulose plates. Another advantage of the cellulose-supported media, that no solidifying aid is needed for solidification regardless of culture conditions, is also discussed.     

不同无机碳源对粉核油球藻生长的影响

研究了2种无机碳源对粉核油球藻(Pinguiococcus pyrenoidosus CCMP2078)生长的影响。结果表明,适量添加NaHCO₃和通CO₂对P.pyrenoidosus CCMP2078的生长均有促进作用,NaHCO₃的添加方式对该藻的生长具有显著差异。一次性添加和分批添加NaHCO₃的优化浓度分别为5mmoL·L^-1和20mmoL·L^-1,后者对数生长末期的细胞密度、干重和比生长速率均最高,分别为8.93×10⁶cells·mL^-1,0.247g·L^-1和0.225dL^-1。通入CO₂的体积分数为0.5%时,最有利于藻细胞的生长,对数生长末期的细胞密度、干重和比生长率分别为3.83×10⁷cells·mL^-1、0.42g·L^-1和0.212d^-1。     

Preferential utilization of organic and inorganic sources of phosphorus by wheat plant

On soils of low P supply organic P (Po) makes up a similar or even larger part in soil solution than inorganic P (Pi). The ability of wheat (Triticum aestivum L., cv. Star) plants to hydrolyze and absorb this Po in comparison to similar concentrations of Pi was studied. Four concentration levels of Pi and Po were obtained by extracting two soils with deionized water in a ratio of 1:1 and concentrating the resulting filtrate by freeze drying to different degrees. The concentration of Pi varied between 5 and 36 μM and Po between 3 and 22 μM. Wheat seedlings were grown in these solutions for 12 and 24 h and acid and alkaline phosphatase activity determined. The reduction of Po concentration in solution expressed on a root length basis gave the rate of Po hydrolysis and the reduction in concentration of Pi and Po gave the P inflow into the roots. No alkaline phosphatase activity was detected. The activity of wheat root acid phosphatase increased with Po concentration in solution. Phosphorus uptake was 2 to 6 fold higher from Pi than from Po at similar concentrations of both. The rate of uptake from Pi, the inflow, as well as the rate of hydrolysis of Po increased linearly with concentration but at similar concentration the inflow was 2 to 4 times higher than the rate of Po hydrolysis. Results suggest that plants can utilize Po after hydrolysis by phosphatase, but Pi is more important and preferentially used by plants; Po may be essential for plant nutrition especially in high P-fixing soils.     

Investigation of mercaptans,organic sulfides,and inorganic sulfur compounds as sulfur sources for the growth of methanogenic bacteria

A variety of compounds were investigated for use as sulfur sources for the growth of methanogenic bacteria.Methanococcus (Mc.) deltae, Mc. maripaludis, Methanobacterium (Mb.) speciesGC-2B, GC-3B, andMMY, Methanobrevibacter (Mbr.) ruminantium, andMethanosarcina (Ms.) barkeri strain 227 grew well with sulfide, S^o, thiosulfate, or cysteine as sole sulfur source.Mbr. ruminatium was able to grow on SO ₄ ⁼ or SO ₃ ⁼ , andMs. barkeri strain 227 was able to grow on SO ₃ ⁼ , but not on SO ₄ ⁼ as a sole sulfur source.Mc. jannaschii grew with sulfide, S^o, thiosulfate or SO ₃ ⁼ , but not on cysteine or SO ₄ ⁼ as sole surface source.Mc. thermolithotrophicus, Mc. jannaschii, Mc. deltae, andMb. thermoautotrophicum strains Marburg and H were able to grow with methanethiol, ethanethiol,n-propanethiol,n-butanethiol, methyl sulfide, dimethyl sulfoxide, ethyl sulfide, or CS₂ as a sulfur source, when very low levels (20–30 M) of sulfide were present; no growth occurred on 5–100 M sulfide alone. Methanethiol, ethanethiol, and methyl sulfide-using cultures produced sulfide during growth.