Crystal Structures of Alkylperoxo and Anhydride Intermediates in an Intradiol Ring-cleaving Dioxygenase


Book Description

Intradiol aromatic ring-cleaving dioxygenases use an active site, nonheme Fe3+ to activate O2 and catecholic substrates for reaction. The inability of Fe3+ to directly bind O2 presents a mechanistic conundrum. The reaction mechanism of protocatechuate 3,4-dioxygenase is investigated in this paper using the alternative substrate 4-fluorocatechol. This substrate is found to slow the reaction at several steps throughout the mechanistic cycle, allowing the intermediates to be detected in solution studies. When the reaction was initiated in an enzyme crystal, it was found to halt at one of two intermediates depending on the pH of the surrounding solution. The X-ray crystal structure of the intermediate at pH 6.5 revealed the key alkylperoxo-Fe3+ species, and the anhydride-Fe3+ intermediate was found for a crystal reacted at pH 8.5. Intermediates of these types have not been structurally characterized for intradiol dioxygenases, and they validate four decades of spectroscopic, kinetic, and computational studies. In contrast to our similar in crystallo crystallographic studies of an Fe2+-containing extradiol dioxygenase, no evidence for a superoxo or peroxo intermediate preceding the alkylperoxo was found. This observation and the lack of spectroscopic evidence for an Fe2+ intermediate that could bind O2 are consistent with concerted formation of the alkylperoxo followed by Criegee rearrangement to yield the anhydride and ultimately ring-opened product. Finally, structural comparison of the alkylperoxo intermediates from the intra- and extradiol dioxygenases provides a rationale for site specificity of ring cleavage.




Iron-containing Enzymes


Book Description

Mononuclear iron containing enzymes are important intermediates in bioprocesses and have potential in the industrial biosynthesis of specific products. This book features topical review chapters by leaders in this field and its various sub-disciplines.










Journal


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Approaches in Bioremediation


Book Description

Bioremediation refers to the clean‐up of pollution in soil, groundwater, surface water, and air using typically microbiological processes. It uses naturally occurring bacteria and fungi or plants to degrade, transform or detoxify hazardous substances to human health or the environment. For bioremediation to be effective, microorganisms must enzymatically attack the pollutants and convert them to harmless products. As bioremediation can be effective only where environmental conditions permit microbial growth and action, its application often involves the management of ecological factors to allow microbial growth and degradation to continue at a faster rate. Like other technologies, bioremediation has its limitations. Some contaminants, such as chlorinated organic or high aromatic hydrocarbons, are resistant to microbial attack. They are degraded either gradually or not at all, hence, it is not easy to envisage the rates of clean-up for bioremediation implementation. Bioremediation represents a field of great expansion due to the important development of new technologies. Among them, several decades on metagenomics expansion has led to the detection of autochthonous microbiota that plays a key role during transformation. Transcriptomic guides us to know the expression of key genes and proteomics allow the characterization of proteins that conduct specific reactions. In this book we show specific technologies applied in bioremediation of main interest for research in the field, with special attention on fungi, which have been poorly studied microorganisms. Finally, new approaches in the field, such as CRISPR-CAS9, are also discussed. Lastly, it introduces management strategies, such as bioremediation application for managing affected environment and bioremediation approaches. Examples of successful bioremediation applications are illustrated in radionuclide entrapment and retardation, soil stabilization and remediation of polycyclic aromatic hydrocarbons, phenols, plastics or fluorinated compounds. Other emerging bioremediation methods include electro bioremediation, microbe-availed phytoremediation, genetic recombinant technologies in enhancing plants in accumulation of inorganic metals, and metalloids as well as degradation of organic pollutants, protein-metabolic engineering to increase bioremediation efficiency, including nanotechnology applications are also discussed.




Plant Microbiomes for Sustainable Agriculture


Book Description

This book encompasses the current knowledge of plant microbiomes and their potential biotechnological application for plant growth, crop yield and soil health for sustainable agriculture. The plant microbiomes (rhizospheric, endophytic and epiphytic) play an important role in plant growth, development, and soil health. Plant and rhizospheric soil are a valuable natural resource harbouring hotspots of microbes, and it plays critical roles in the maintenance of global nutrient balance and ecosystem function. The diverse group of microbes is key components of soil–plant systems, where they are engaged in an intense network of interactions in the rhizosphere/endophytic/phyllospheric. The rhizospheric microbial diversity present in rhizospheric zones has a sufficient amount of nutrients release by plant root systems in form of root exudates for growth, development and activities of microbes. The endophytic microbes are referred to those microorganisms, which colonize in the interior of the plant parts, viz root, stem or seeds without causing any harmful effect on host plant. Endophytic microbes enter in host plants mainly through wounds, naturally occurring as a result of plant growth, or through root hairs and at epidermal conjunctions. Endophytes may be transmitted either vertically (directly from parent to offspring) or horizontally (among individuals). The phyllosphere is a common niche for synergism between microbes and plant. The leaf surface has been termed as phyllosphere and zone of leaves inhabited by microorganisms as phyllosphere. The plant part, especially leaves, is exposed to dust and air currents resulting in the establishments of typical flora on their surface aided by the cuticles, waxes and appendages, which help in the anchorage of microorganisms. The phyllospheric microbes may survive or proliferate on leaves depending on extent of influences of material in leaf diffuseness or exudates. The leaf diffuseness contains the principal nutrients factors (amino acids, glucose, fructose and sucrose), and such specialized habitats may provide niche for nitrogen fixation and secretions of substances capable of promoting the growth of plants. The microbes associated with plant as rhizospheric, endophytic and epiphytic with plant growth promoting (PGP) attributes have emerged as an important and promising tool for sustainable agriculture. PGP microbes promote plant growth directly or indirectly, either by releasing plant growth regulators; solubilization of phosphorus, potassium and zinc; biological nitrogen fixation or by producing siderophore, ammonia, HCN and other secondary metabolites which are antagonistic against pathogenic microbes. The PGP microbes belong to different phylum of archaea (Euryarchaeota); bacteria (Acidobacteria, Actinobacteria, Bacteroidetes, Deinococcus-Thermus, Firmicutes and Proteobacteria) and fungi (Ascomycota and Basidiomycota), which include different genera namely Achromobacter, Arthrobacter, Aspergillus, Azospirillum, Azotobacter, Bacillus, Beijerinckia, Burkholderia, Enterobacter, Erwinia, Flavobacterium, Gluconoacetobacter, Haloarcula, Herbaspirillum, Methylobacterium, Paenibacillus, Pantoea, Penicillium, Piriformospora, Planomonospora, Pseudomonas, Rhizobium, Serratia and Streptomyces. These PGP microbes could be used as biofertilizers/bioinoculants at place of chemical fertilizers for sustainable agriculture. The aim of “Plant Microbiomes for Sustainable Agriculture” is to provide the current developments in the understanding of microbial diversity associated with plant systems in the form of rhizospheric, endophytic and epiphytic. The book is useful to scientist, research and students related to microbiology, biotechnology, agriculture, molecular biology, environmental biology and related subjects.