Abstract:The main objective of the immobilization of enzymes is to enhance the economics of biocatalytic processes. Immobilization allows one to re-use the enzyme for an extended period of time and enables easier separation of the catalyst from the product. Additionally, immobilization improves many properties of enzymes such as performance in organic solvents, pH tolerance, heat stability or the functional stability. Increasing the structural rigidity of the protein and stabilization of multimeric enzymes which prevents dissociation-related inactivation. In the last decade, several papers about immobilization methods have been published. In our work, we present a relation between the influence of immobilization on the improvement of the properties of selected oxidoreductases and their commercial value. We also present our view on the role that different immobilization methods play in the reduction of enzyme inhibition during biotechnological processes.

Currently, analgesics and nonsteroidal anti-inflammatory drugs (NSAIDs) are classified as one of the most emerging group of xenobiotics and have been detected in various natural matrices. Among them, monocyclic paracetamol and ibuprofen, widely used to treat mild and moderate pain are the most popular. Since long-term adverse effects of these xenobiotics and their biological and pharmacokinetic activity especially at environmentally relevant concentrations are better understood, degradation of such contaminants has become a major concern. Moreover, to date, conventional wastewater treatment plants (WWTPs) are not fully adapted to remove that kind of micropollutants. Bioremediation processes, which utilize bacterial strains with increased degradation abilities, seem to be a promising alternative to the chemical methods used so far. Nevertheless, despite the wide prevalence of paracetamol and ibuprofen in the environment, toxicity and mechanism of their microbial degradation as well as genetic background of these processes remain not fully characterized. In this review, we described the current state of knowledge about toxicity and biodegradation mechanisms of paracetamol and ibuprofen and provided bioinformatics analysis concerning the genetic bases of these xenobiotics decomposition.

Endophytic bacteria, which interact closely with their host, are an essential part of the plant microbiome. These interactions enhance plant tolerance to environmental changes as well as promote plant growth, thus they have become attractive targets for increasing crop production. Numerous studies have aimed to characterise how endophytic bacteria infect and colonise their hosts as well as conferring important traits to the plant. In this review, we summarise the current knowledge regarding endophytic colonisation and focus on the insights that have been obtained from the mutants of bacteria and plants as well as ‘omic analyses. These show how endophytic bacteria produce various molecules and have a range of activities related to chemotaxis, motility, adhesion, bacterial cell wall properties, secretion, regulating transcription and utilising a substrate in order to establish a successful interaction. Colonisation is mediated by plant receptors and is regulated by the signalling that is connected with phytohormones such as auxin and jasmonic (JA) and salicylic acids (SA). We also highlight changes in the expression of small RNAs and modifications of the cell wall properties. Moreover, in order to exploit the beneficial plant-endophytic bacteria interactions in agriculture successfully, we show that the key aspects that govern successful interactions remain to be defined.

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Stenotrophomonas maltophilia KB2 used in this study is known to metabolise broad range of aromatic compounds including phenol, some chloro and methylphenols, benzoic acids, catochols and others. To study the applicability of the strain for degradation of mononitrophenols in monosubstrate as well as cometabolic systems its degradation potential in the presence of mononitrophenols or different aromatic compounds of plant origin was tested. Stenotrophomonas maltophilia KB2 strain was not able to degrade any of mononitrophenols used in the single substrate experiments. Effect of additional carbon source on nitrophenols degradation revealed that presence of benzoate, 4-hydroxybenzoate or 3,4-dixydroxybenzoate stimulate transformation of 2-nitrophenol, 3-nitrophenol as well as 4-nitrophenol. Depending on growth substrate and mononitrophenol used, decrease in cometabolite concentration was from 25 to 45%. Obtained results suggest that Stenotrophomonas maltophilia KB2 strain could be potentially used for cometabolic degradation of nitrophenols in the presence of aromatic acids, for the bioremediation of contaminated sites.

Stenotrophomonas maltophilia KB2 is known to produce different enzymes of dioxygenase family. The aim of our studies was to determine activity of these enzymes after induction by benzoic acids in cometabolic systems with nitrophenols. We have shown that under cometabolic conditions KB2 strain degraded 0.25–0.4 mM of nitrophenols after 14 days of incubation. Simultaneously degradation of 3 mM of growth substrate during 1–3 days was observed depending on substrate as well as cometabolite used. From cometabolic systems with nitrophenols as cometabolites and 3,4-dihydroxybenzoate as a growth substrate, dioxygenases with the highest activity of protocatechuate 3,4-dioxygenase were isolated. Activity of catechol 1,2- dioxygenase and protocatechuate 4,5-dioxygenase was not observed. Catechol 2,3-dioxygenase was active only in cultures with 4-nitrophenol. Ability of KB2 strain to induce and synthesize various dioxygenases depending on substrate present in medium makes this strain useful in bioremediation of sites contaminated with different aromatic compounds.

This is the first report of a catechol 1,2-dioxygenase from Stenotrophomonas maltophilia strain KB2 with high activity against catechol and its methyl derivatives. This enzyme was maximally active at pH 8.0 and 40 °C and the half-life of the enzyme at this temperature was 3 h. Kinetic studies showed that the value of Km and Vmax was 12.8 μM and 1,218.8 U/mg of protein, respectively. During our studies on kinetic properties of the catechol 1,2-dioxygenase we observed substrate inhibition at >80 μM. The nucleotide sequence of the gene encoding the S. maltophilia strain KB2 catechol 1,2-dioxygenase has high identity with other catA genes from members of the genus Pseudomonas. The deduced 314-residue sequence of the enzyme corresponds to a protein of molecular mass 34.5 kDa. This enzyme was inhibited by competitive inhibitors (phenol derivatives) only by ca. 30 %. High tolerance against condition changes is desirable in industrial processes. Our data suggest that this enzyme could be of use as a tool in production of cis,cis-muconic acid and its derivatives.