Genomic analysis of the wheat rhizosphere-isolated Streptomyces acrimycini Encodes Enzymes with Potential Applications in Biotechnology

Chapter 11 - Exploration of microbial communities of Indian hot springs and their potential biotechnological applications

Study the effect of F17S mutation on the chimeric Bacillus thermocatenulatus lipase

Metabolic flux analysis is a vital tool used to determine the ultimate output of cellular metabolism and thus detect biotechnologically relevant bottlenecks in productivity. 13C-based metabolic flux analysis (13C-MFA) and flux balance analysis (FBA) have many potential applications in biotechnology. However, noteworthy hurdles in fluxomics study are still present. First, several technical difficulties in both 13C-MFA and FBA severely limit the scope of fluxomics findings and the applicability of obtained metabolic information. Second, the complexity of metabolic regulation poses a great challenge for precise prediction and analysis of metabolic networks, as there are gaps between fluxomics results and other omics studies. Third, despite identified metabolic bottlenecks or sources of host stress from product synthesis, it remains difficult to overcome inherent metabolic robustness or to efficiently import and express nonnative pathways. Fourth, product yields often decrease as the number of enzymatic steps increases. Such decrease in yield may not be caused by rate-limiting enzymes, but rather is accumulated through each enzymatic reaction. Fifth, a high-throughput fluxomics tool hasnot been developed for characterizing nonmodel microorganisms and maximizing their application in industrial biotechnology. Refining fluxomics tools and understanding these obstacles will improve our ability to engineer highlyefficient metabolic pathways in microbial hosts.

The combination of magnetic and luminescent properties in a single particle system, opens-up a wide range of potential applications in biotechnology and biomedicine. In this work, we performed the synthesis of magnetic-luminescent Gd2O3:Eu3+@Fe2O3 particles by ultrasonic spray pyrolysis performed in a tubular furnace. In order to achieve the composite formation, commercial superparamagnetic Fe3O4 nanoparticles were coated with a luminescent Eu3+-doped Gd2O3 shell in a low-cost one-step process. The spray pyrolysis method yields deagglomerated spherical shape magneto/luminescent particles. The photoluminescence spectra under UV excitation (λExc = 265 nm) of the magnetic Gd2O3:Eu3+@Fe2O3 compound showed the characteristic red emission of Eu3+ (λEm = 612 nm). This magneto/luminescent system will find applications in biomedicine and biotechnology.

Ion channels in key marine invertebrates; their diversity and potential for applications in biotechnology

The RNA phages offer promising applications in biotechnology, including vaccine development and drug delivery. However, their potential remains underexplored due to the limited number of known RNA phages, partly because conventional methods fail to identify plaque-invisible lytic phages that do not form plaques. Here, we introduced a novel method that combines RNA-inclusive metagenomic studies and quantitative reverse transcription-PCR (RMS-RT-qPCR) to identify and characterize active RNA phages from environmental samples. This study led to the discovery of a new active Qbeta-like phage, named Cute. Genomic analysis revealed that Cute is a new member of the Qubevirus genus. Although Cute does not form plaques, it can be observed to continuously release into the supernatant when co-cultured with the host by RT-qPCR detection. This discovery underscores the potential diversity of RNA phages in nature and the limitations of traditional culture-dependent techniques. Our findings suggest that RMS-RT-qPCR could aid in the discovery of active RNA phages with significant biotechnological applications.IMPORTANCEThe discovery and characterization of RNA phages might be historically constrained by traditional culture-based methods. Our study provides a powerful tool for identifying active RNA phages by combining RNA-inclusive metagenomic analysis with RT-qPCR. This method expands our understanding of the diversity and ecological roles of RNA phages, which are often overlooked in microbiome studies. This research highlights the importance of RNA phages in natural ecosystems and their potential applications in biotechnology and medicine, such as antimicrobial therapies and vaccine development. By expanding our understanding of RNA phage diversity, this study opens new avenues for their utilization in various fields, emphasizing the need for continued exploration of these versatile biological entities.

Metagenome-assembled genomes (MAGs) have contributed to identifying non-culturable microorganisms and understanding their ecological functions. MAGs offer an advantage in investigating sporulation-associated genes, especially given the difficulty of isolating many species residing in the gut microbiota of multiple hosts. Bacterial sporulation is a key survival mechanism with implications for pathogenicity and biotechnology. Here, we investigate MAGs from vertebrate hosts, emphasizing taxonomic identification and identifying sporulation-associated genes in potential novel species within the Firmicutes phylum. We identified potential new species in the classes Clostridia (Borkfalkiaceae, Lachnospiraceae, Monoglobaceae, and Oscillospiraceae families) and Bacilli (Bacillaceae and Erysipelotrichaceae families) through phylogenetic and functional pathway analyses, highlighting their sporulation potential. Our study covers 146 MAGs, 124 of them without refined taxonomic assignments at the family level. We found that Clostridia and Bacilli have unique sporulation gene profiles in the refined family MAGs for cattle, swine, poultry, and human hosts. The presence of genes related to Spo0A regulon, engulfment, and spore cortex in MAGs underscores fundamental mechanisms in sporulation processes in currently uncharacterized species with sporulation potential from metagenomic dark matter. Furthermore, genomic analyses predict sporulation potential based on gene presence, genome size, and metabolic pathways involved in spore formation. We emphasize MAGs covering families not yet characterized through the phylogenetic analysis, and with extensive potential for spore-forming bacteria within Clostridia, Bacilli, UBA4882, and UBA994 classes. These findings contribute to exploring spore-forming bacteria, which provides evidence for novel species diversity in multiple hosts, their adaptive strategies, and potential applications in biotechnology and host health.IMPORTANCESpores are essential for bacterial survival in harsh environments, facilitating their persistence and adaptation. Exploring sporulation-associated genes in metagenome-assembled genomes (MAGs) from different hosts contributes to clinical and biotechnological domains. Our study investigated the extent of genes associated with bacterial sporulation in MAGs from poultry, swine, cattle, and humans, revealing these genes in uncultivated bacteria. We identified potential novel Firmicutes species with sporulation capabilities through phylogenetic and functional analyses. Notably, MAGs belonging to Clostridia, Bacilli, and unknown classes, namely UBA4882 and UBA994, remained uncharacterized at the family level, which raises the hypothesis that sporulation would also be present in these genomes. These findings contribute to our understanding of microbial adaptation and have implications for microbial ecology, underlining the importance of sporulation in Firmicutes across different hosts. Further studies into novel species and their sporulation capability can contribute to bacterial maintenance mechanisms in various organisms and their applications in biotechnology studies.

Toxins purified from the venom of spiders have high potential to be studied pharmacologically and biochemically. These biomolecules may have biotechnological and therapeutic applications. This study aimed to evaluate the protein content of Parawixia bistriata venom and functionally characterize its proteins that have potential for biotechnological applications. The crude venom showed no phospholipase, hemorrhagic, or anti-Leishmania activities attesting to low genotoxicity and discrete antifungal activity for C. albicans. However the following activities were observed: anticoagulation, edema, myotoxicity and proteolysis on casein, azo-collagen, and fibrinogen. The chromatographic and electrophoretic profiles of the proteins revealed a predominance of acidic, neutral, and polar proteins, highlighting the presence of proteins with high molecular masses. Five fractions were collected using cation exchange chromatography, with the P4 fraction standing out as that of the highest purity. All fractions showed proteolytic activity. The crude venom and fractions P1, P2, and P3 showed larvicidal effects on A. aegypti. Fraction P4 showed the presence of a possible metalloprotease (60 kDa) that has high proteolytic activity on azo-collagen and was inhibited by EDTA. The results presented in this study demonstrate the presence of proteins in the venom of P. bistriata with potential for biotechnological applications.

Steroid compounds are produced by eukaryotes where they have a variety of chemical structures and play important physiological roles. Many bacteria are capable of transforming and completely degrading steroids under various growth conditions. The microbial metabolism of steroids has gained considerable interest due to its potential applications in industrial and environmental biotechnology. The oxic degradation pathways of steroids and some of the involved enzymes are well characterized. The key players in these pathways are oxygenases which depend on dioxygen as a co-substrate. On the contrary, much less is known about the mechanisms operating under anoxic conditions. Obviously, anoxic bacterial metabolism of steroids should proceed via oxygenase-independent reactions. So far, a few bacteria that can completely degrade steroids in the absence of oxygen were characterized. Surprisingly, all of them belong to denitrifying bacteria and utilize only nitrate as the alternative electron acceptor. Recent studies of anoxic metabolism of steroids using denitrifying bacteria revealed unique and interesting biochemical reactions and enzymes. Here we discuss the current understanding of the biochemistry and molecular biology of bacterial steroid metabolism under anoxic conditions. The aerobic metabolism of steroids is briefly presented for the sake of comparison. Future investigations on anoxic metabolism of steroids will unravel novel aspects of the regulation and evolution of catabolic pathways as well as unprecedented biocatalysts with useful applications in biotechnology.

α-Amylases (1,4-α-D-glucan-4-glucanohydrolase, EC 3.2.1.1) are endoamylases catalyzing the hydrolysis of α- (1→4) glucosidic linkages of polysaccharides such as starch, glycogen etc. In general they belong to class-13 of glycosyl hydrolases. α-Amylases have the major world market share of enzymes. These endoamylases are one of the most important enzymes for industrial and biotechnological applications. It holds key application in detergent, food, textile, pharmaceuticals, and bakery industries. In this review, we summarize some important features, mechanism and physicochemical properties of α-amylase family. In addition, applications and future prospects of industrially important enzyme having potential for biotechnological applications are discussed in detail. Keywords: α-Amylases, Amylases, glycosyhydrolase, starch, structure, calcium, thermostability, biosensors, disulphide bond, single polypeptide chains, isoform AMY1, gibberellic acid, abscisic acid, Biosynthesis, dextrins, glucose, applications

This paper investigates the adsorption of bovine serum albumin (BSA) and bovine hemoglobin (BHb) model proteins onto novel thymine-functionalized polystyrene (PS-VBT) microspheres, in comparison with polystyrene (PS) microspheres. Maximum adsorption was obtained for both proteins near their corresponding isoelectric points (pI at pH = 4.7 for BSA and 7.1 for BHb). FTIR and adsorption isotherm analysis demonstrated that, although both proteins were physisorbed onto PS through nonspecific hydrophobic interactions, adsorption onto the functionalized copolymers occurred by both physisorption and chemisorption via hydrogen bonding. FTIR analysis also indicated conformational changes in the secondary structure of BSA and BHb adsorbed onto PS, whereas little or no conformation change was seen in the case of adsorption onto PS-VBT. Atomic force microscopy (AFM), consistent with the isotherm results, also demonstrated monolayer adsorption for both proteins. AFM images of BSA adsorbed onto copolymers with 20 mol % surface VBT loading showed exclusively end-on orientation. Adsorption onto copolymers with lower functionality showed mixed end-on and side-on orientation modes of BSA, and only the side-on orientation was observed on PS. The AFM results agreed well with theoretically calculated and experimentally obtained adsorption capacities. AFM together with calculated and observed adsorption capacity data for BHb indicated that this protein might be highly compressed on the copolymer surface. Adsorption from a binary mixture of BSA and BHb onto PS-VBT showed good separation at pH=7.0; approximately 90% of the adsorbed protein was BHb. The novel copolymers have potential applications in biotechnology.

Ice-binding proteins (IBPs), such as antifreeze proteins (AFPs) and ice-nucleating proteins (INPs), have been described in diverse cold-adapted organisms, and their potential applications in biotechnology have been recognized in various fields. Currently, both IBPs are being applied to biotechnological processes, primarily in medicine and the food industry. However, our knowledge regarding the diversity of bacterial IBPs is limited; few studies have purified and characterized AFPs and INPs from bacteria. Phenotypically verified IBPs have been described in members belonging to Gammaproteobacteria, Actinobacteria and Flavobacteriia classes, whereas putative IBPs have been found in Gammaproteobacteria, Alphaproteobacteria and Bacilli classes. Thus, the main goal of this minireview is to summarize the current information on bacterial IBPs and their application in biotechnology, emphasizing the potential application in less explored fields such as agriculture. Investigations have suggested the use of INP-producing bacteria antagonists and AFPs-producing bacteria (or their AFPs) as a very attractive strategy to prevent frost damages in crops. UniProt database analyses of reported IBPs (phenotypically verified) and putative IBPs also show the limited information available on bacterial IBPs and indicate that major studies are required.

Psychrophilic bacteria are a large group of microorganisms that prevail in low-temperature ecosystems. Psychrophilic bacteria have undergone a number of adaptations that help them exist in such conditions. One of such adaptations is the use of enzymes with a high specific activity at low temperatures. Such enzymes are usually called “cold-active.” These enzymes have potential applications in biotechnology and industry. In our review, we considered individual genera of psychrophilic bacteria, current global trends in the study of cold-active enzymes, their applications, and place in industrial biotechnology. Thus, the main goal of this study was to explore the diversity of psychrophilic bacteria, as well as opportunities of their application in biotechnology. The natural ecological sites of psychrophiles are numerous and varied. Psychrophiles form a permanent microflora of eternal cold regions, polar regions and oceans. Bacteria belonging to this group are found in soil, water or associated with plants and animals. An important site for psychrophilic microorganisms is a low-temperature water reservoir. At present, many new genera of psychrophiles and psychrotrophs have been derived from the bottom sediments and sea waters of the Arctic and Antarctic and described. Psychrophilic microorganisms are found in caves and in ancient ice crystal structures. The latter testifies to the very possibility of the super-long anabiosis phenomenon, as well as vital capacity preservation without division for a long period of time. Psychrophiles do not have a single form, they belong to at least several phylogenetic groups. Psychrophilic forms are found among the representatives of a large number of genera. There are no common physiological and biochemical parameters typical of psychrophilic bacteria. They comprise rods, cocci, vibrios, gram-negative and gram-positive bacteria, bacteria that produce and do not produce spores, strict aerobes, facultative and strict anaerobes. We lay greater emphasis on the diversity of psychrophilic bacteria capable of producing industrially important enzymes. The review considers bacteria belonging to the genera Vibrio and Aliivibrio, Pseudomonas, Achromobacter, Arthrobacter, Pseudoalteromonas, Bacillus, Clostridium, Micrococcus, Psychrobacter, Psychromonas, Flavobacterium, and psychrophilic methanotrophic microorganisms. These bacteria enzymes are used in agriculture, biotechnology, pharmaceuticals and household chemicals, as well as other sectors of the national economy. Psychrophilic bacteria produce a chemical compound that can be used in medicine. For example, Pseudomonas antarctica contains a cluster of genes encoding microcin B, R-type pyocins, adenosylcobalamin, and pyrroloquinoline quinone. Thus, P. antarctica has antibiotic activity. Psychrobacter proteolyticus also has an antineoplastic action and secrets an extracellular cold-adapted metalloproteinase being able to inhibit the space-occupying process. Cold-active metalloproteinases are also widely used as detergents, in currying, food sector and molecular biology. The immunogenic Pal conformable protein was derived from the psychrophilic strain of The representatives of the genus Arthrobacter capable of metabolizing diuron and petroleum products have an important property. A. agilis produces a red pigment, a bacterioruberin-type carotenoid being interesting as an antioxidant. A. psychrochitiniphilus is promising for cleaning water areas, oil-polluted coastlines, as it decomposes oil and petroleum products. Flavobacterium limicola is a potential source of cold-active protease. This bacterium is characterized by an increase in protease secretion as temperature decreases. Thus, F. limicola can be used in environmental biotransformations and bioremediations. The psychrophilic bacteria of the genus Bacillus are the participants of active studies. Their cold-active enzymes have a high potential in various areas of biomedicine, immunology, decontamination, and various industrial applications. The antifreeze proteins of psychrophilic Clostridia are considered a promising biotechnological product for use in medicine, food, beauty products, fuel, and other industries. This study reviews literary sources and indicates that at present obligate and facultative psychrophiles (psychrotrophs) and their cold-active enzymes are of scientific interest throughout the world. A significant part of the research is focused on a general understanding of the distribution of psychrophilic bacteria and a local study of enzymatic activity. A further study of psychrophilic microorganisms producing enzymes at low temperatures will reveal new ways for the development of biotechnologies in various sectors of the national economy. The paper contains 94 References. The Authors declare no conflict of interest.

In this article, we report an efficient method for the synthesis of thymine‐functionalized polystyrene microspheres. First, poly(styrene‐co‐4‐chloromethylstyrene) copolymers slightly crosslinked with divinylbenzene were synthesized in batch free‐radical emulsion copolymerization. Microspheres with a particle size of ∼40–70 nm were obtained with greater than 99% conversion. The chloromethylstyrene (CMS) groups were then converted into thymylmethylstyrene (TMS) in a two‐phase system with greater than 80% efficiency, and up to a 45 mol % thymine loading was achieved. The functionalized microspheres were characterized by elemental analysis, Fourier transform infrared, and X‐ray photoelectron spectroscopy. The analyses revealed partial hydrolysis of the CMS functionalities, yielding hydroxymethyl functional groups in addition to the thymine functionalities. These copolymers have potential applications in biotechnology. © 2005 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 43: 5545–5553, 2005

Hypersaline environments as natural sources of microbes with potential applications in biotechnology: The case of solar evaporation systems to produce salt in Alicante County (Spain).