Polyphosphate Kinase from , One Enzyme Catalyzing a Two-Step Cascade Reaction to Synthesize ATP from AMP.

This study characterizes a novel polyphosphate kinase from (PPK2-III), an enzyme with potential applications in ATP regeneration processes. Bioinformatic and structural analyses confirmed the presence of conserved motifs characteristic of PPK2 enzymes, including Walker A and B motifs, and the subclass-specific residue E137. Molecular docking simulations showed AMP had the highest binding affinity (-7.

Lysozyme-Responsive Hydrogels of Chitosan-Streptomycin Conjugates for the On-Demand Release of Biofilm-Dispersing Enzymes for the Efficient Eradication of Oral Biofilms.

Hydrogels with controlled degradation and sustained antibiofilm activity are promising biomaterials for the treatment of oral infections such as periodontitis or caries. In this article, an forming chitosan-streptomycin hydrogel is developed that can target established bacterial biofilms in response to lysozyme, an enzyme that is overexpressed in saliva during oral infections. When the new hydrogel is applied to simulated oral biofilms, the overexpressed lysozyme degrades the hydrogel and releases chitosan-streptomycin oligosaccharides that can eradicate the biofilm.

Synthesis of Chiral Acyclic Pyrimidine Nucleoside Analogues from DHAP-Dependent Aldolases.

Dihydroxyacetone phosphate (DHAP)-dependent aldolases catalyze the aldol addition of DHAP to a variety of aldehydes and generate compounds with two stereocenters. This reaction is useful to synthesize chiral acyclic nucleosides, which constitute a well-known class of antiviral drugs currently used. In such compounds, the chirality of the aliphatic chain, which mimics the open pentose residue, is crucial for activity.

From the problem to the solution: Chitosan valorization cycle.

The problem of fisheries waste has increased in recent years and has become a global problem influenced by various biological, technical, operational and socioeconomic factors. In this context, the use of these residues as raw materials is a proven approach not only to reduce the crisis of unprecedented magnitude facing the oceans, but also to improve the management of marine resources and increase the competitiveness of the fisheries sector. However, the implementation of valorization strategies at the industrial level is being excessively slow, despite this great potential.

Fast and Sustained Axonal Growth by BDNF Released from Chitosan Microspheres.

Brain-derived neurotrophic factor (BDNF) regulates dendritic branching and dendritic spine morphology, as well as synaptic plasticity and long-term potentiation. Consequently, BDNF deficiency has been associated with some neurological disorders such as Alzheimer's, Parkinson's or Huntington's diseases. In contrast, elevated BDNF levels correlate with recovery after traumatic central nervous system (CNS) injuries.

Design of a biocatalytic cascade for the enzymatic sulfation of unsulfated chondroitin with generation of PAPS.

Sulfation of molecules in living organisms is a process that plays a key role in their functionality. In mammals, the sulfation of polysaccharides (glycosaminoglycans) that form the proteoglycans present in the extracellular matrix is particularly important. These polysaccharides, through their degree and sulfation pattern, are involved in a variety of biological events as signal modulators in communication processes between the cell and its environment.

Design and biocatalytic applications of genetically fused multifunctional enzymes.

Fusion proteins, understood as those created by joining two or more genes that originally encoded independent proteins, have numerous applications in biotechnology, from analytical methods to metabolic engineering. The use of fusion enzymes in biocatalysis may be even more interesting due to the physical connection of enzymes catalyzing successive reactions into covalently linked complexes. The proximity of the active sites of two enzymes in multi-enzyme complexes can make a significant contribution to the catalytic efficiency of the reaction.

Chitosan sulfate-lysozyme hybrid hydrogels as platforms with fine-tuned degradability and sustained inherent antibiotic and antioxidant activities.

The control of the properties and biological activities of chitosan-lysozyme hybrid hydrogels to exploit their interesting biomedical applications depends largely on the chitosan acetylation pattern, a difficult parameter to control. Herein, we have prepared sulfated chitosan-lysozyme hydrogels as versatile platforms with fine-tuned degradability and persistent bactericidal and antioxidant properties. The use of chitosan sulfates instead of chitosan has the advantage that the rate and mechanisms of lysozyme release, as well as antibacterial and antioxidant activities, depend on the sulfation profile, a structural parameter that is easily controlled by simple chemical modifications.

Preparation and Characterization of Aminoglycoside-Loaded Chitosan/Tripolyphosphate/Alginate Microspheres against .

Although aminoglycosides are one of the common classes of antibiotics that have been widely used for treating infections caused by pathogenic bacteria, the evolution of bacterial resistance mechanisms and their inherent toxicity have diminished their applicability. Biocompatible carrier systems can help sustain and control the delivery of antibacterial compounds while reducing the chances of antibacterial resistance or accumulation in unwanted tissues. In this study, novel chitosan gel beads were synthesized by a double ionic co-crosslinking mechanism.

Heparanized chitosans: towards the third generation of chitinous biomaterials.

The functionalization of chitosans is an emerging research area in the design of solutions for a wide range of biomedical applications. In particular, the modification of chitosans to incorporate sulfate groups has generated great interest since they show structural similarity to heparin and heparan sulfates. Most of the biomedical applications of heparan sulfates are derived from their ability to bind different growth factors and other proteins, as through these interactions they can modulate the cellular response.

Deciphering Structural Determinants in Chondroitin Sulfate Binding to FGF-2: Paving the Way to Enhanced Predictability of their Biological Functions.

Controlling chondroitin sulfates (CSs) biological functions to exploit their interesting potential biomedical applications requires a comprehensive understanding of how the specific sulfate distribution along the polysaccharide backbone can impact in their biological activities, a still challenging issue. To this aim, herein, we have applied an "holistic approach" recently developed by us to look globally how a specific sulfate distribution within CS disaccharide epitopes can direct the binding of these polysaccharides to growth factors. To do this, we have analyzed several polysaccharides of marine origin and semi-synthetic polysaccharides, the latter to isolate the structure-activity relationships of their rare, and even unnatural, sulfated disaccharide epitopes.

Unraveling the Structural Landscape of Chitosan-Based Heparan Sulfate Mimics Binding to Growth Factors: Deciphering Structural Determinants for Optimal Activity.

Chitosan sulfates have demonstrated the ability to mimic heparan sulfate (HS) function. In this context, it is crucial to understand how the specific structural properties of HS domains determine their functionalities and biological activities. In this study, several HS-mimicking chitosans have been prepared to mimic the structure of HS domains that have proved to be functionally significant in cell processes.

Simple and Practical Multigram Synthesis of d-Xylonate Using a Recombinant Xylose Dehydrogenase.

An efficient multienzyme system for the preparative synthesis of d-xylonate, a chemical with versatile industrial applications, is described. The multienzyme system is based on d-xylose oxidation catalyzed by the xylose dehydrogenase from and the use of catalytic amounts of NAD. The cofactor is regenerated in situ by coupling the reduction of acetaldehyde into ethanol catalyzed by alcohol dehydrogenase from .

Assembly of glycoamino acid building blocks: a new strategy for the straightforward synthesis of heparan sulfate mimics.

A new strategy that enables a modular straightforward synthesis of heparan sulfate oligosaccharide mimics by the assembly of simple glycoamino acid building blocks is described. The coupling between units is readily carried out by an amidation reaction. Several glycoamino acid oligomers were prepared and their interaction with the FGF2 protein was analyzed.

A holistic approach to unravelling chondroitin sulfation: Correlations between surface charge, structure and binding to growth factors.

Chondroitin sulfate (CS) is a relevant family of polysaccharides that participates in a large variety of biological events that are related to neural processes by regulating various growth factors through the pattern and degree of sulfation of the polysaccharide. However, their own complexity makes their optimization for biomedical applications a difficult undertaking. Thus, a different perspective has to be taken.

Biochemical profiling of rat embryonic stem cells grown on electrospun polyester fibers using synchrotron infrared microspectroscopy.

Therapeutic options for spinal cord injuries are severely limited; current treatments only offer symptomatic relief and rehabilitation focused on educating the individual on how to adapt to their new situation to make best possible use of their remaining function. Thus, new approaches are needed, and interest in the development of effective strategies to promote the repair of neural tracts in the central nervous system inspired us to prepare functional and highly anisotropic polymer scaffolds. In this work, an initial assessment of the behavior of rat neural progenitor cells (NPCs) seeded on poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) fiber scaffolds using synchrotron-based infrared microspectroscopy (SIRMS) is described.

Synthesis, physicochemical characterization and biological evaluation of chitosan sulfate as heparan sulfate mimics.

Despite the relevant biological functions of heparan sulfate (HS) glycosaminoglycans, their limited availability and the chemical heterogeneity from natural sources hamper their use for biomedical applications. Chitosan sulfates (ChS) exhibit structural similarity to HSs and may mimic their biological functions. We prepared a variety of ChS with different degree of sulfation to evaluate their ability to mimic HS in protein binding and to promote neural cell division and differentiation.

Analytical Validation of a New Enzymatic and Automatable Method for d-Xylose Measurement in Human Urine Samples.

Hypolactasia, or intestinal lactase deficiency, affects more than half of the world population. Currently, xylose quantification in urine after gaxilose oral administration for the noninvasive diagnosis of hypolactasia is performed with the hand-operated nonautomatable phloroglucinol reaction. This work demonstrates that a new enzymatic xylose quantification method, based on the activity of xylose dehydrogenase from , represents an excellent alternative to the manual phloroglucinol reaction.

Development of a new method for d-xylose detection and quantification in urine, based on the use of recombinant xylose dehydrogenase from Caulobacter crescentus.

The gene xylB from Caulobacter crescentus has been cloned and expressed in Escherichia coli providing a high yield of xylose dehydrogenase (XylB) production and excellent purity (97%). Purified recombinant XylB showed an absolute dependence on the cofactor NAD(+) and a strong preference for d-xylose against other assayed mono and disaccharides. Additionally, XylB showed strong stability when stored as freeze-dried powder at least 250days both at 4°C and room temperature.

Tuning the Phosphoryl Donor Specificity of Dihydroxyacetone Kinase from ATP to Inorganic Polyphosphate. An Insight from Computational Studies.

Dihydroxyacetone (DHA) kinase from Citrobacter freundii provides an easy entry for the preparation of DHA phosphate; a very important C3 building block in nature. To modify the phosphoryl donor specificity of this enzyme from ATP to inorganic polyphosphate (poly-P); a directed evolution program has been initiated. In the first cycle of evolution, the native enzyme was subjected to one round of error-prone PCR (EP-PCR) followed directly (without selection) by a round of DNA shuffling.

Hyperthermophilic aldolases as biocatalyst for C-C bond formation: rhamnulose 1-phosphate aldolase from Thermotoga maritima.

The TM1072 gene from Thermotoga maritima codifies for a putative form of a rhamnulose-1-phosphate aldolase (Rha-1PA Tm). To investigate this enzyme further, its gene was cloned and expressed in Escherichia coli. The purified enzyme was activated by Co(2+) as a divalent metal ion cofactor, instead of Zn(2+) as its E.

Enzyme catalysed tandem reactions.

To transfer to the laboratory, the excellent efficiency shown by enzymes in Nature, biocatalysis, had to mimic several synthetic strategies used by the living organisms. Biosynthetic pathways are examples of tandem catalysis and may be assimilated in the biocatalysis field for the use of isolated multi-enzyme systems in the homogeneous phase. The concurrent action of several enzymes that work sequentially presents extraordinary advantages from the synthetic point of view, since it permits a reversible process to become irreversible, to shift the equilibrium reaction in such a way that enantiopure compounds can be obtained from prochiral or racemic substrates, reduce or eliminate problems due to product inhibition or prevent the shortage of substrates by dilution or degradation in the bulk media, etc.

Synthesis and evaluation of xylopyranoside derivatives as "decoy acceptors" of human β-1,4-galactosyltransferase 7.

Proteoglycans (PGs), including heparan sulfate forms, are important regulators of tumor progression. In the PGs biosynthetic process, the core protein is synthesized on a ribosomal template and the sugar chains are assembled post-translationally, one sugar at a time, starting with the linkage of xylose to a serine residue of the core protein and followed by galactosidation of the xylosylprotein. Hydrophobic xylopyranosides have been previously shown to prime heparan sulfate synthesis, a property that was required to cause growth inhibition of tumor cells.

Preparation and characterization of a bifunctional aldolase/kinase enzyme: a more efficient biocatalyst for C-C bond formation.

A bifunctional aldolase/kinase enzyme named DLF has been constructed by gene fusion through overlap extension. This fusion enzyme consists of monomeric fructose-1,6-bisphosphate aldolase (FBPA) from Staphylococcus carnosus and the homodimeric dihydroxyacetone kinase (DHAK) from Citrobacter freundii CECT 4626 with an intervening linker of five amino acid residues. The fusion protein was expressed soluble and retained both kinase and aldolase activities.

Substrate channelling in an engineered bifunctional aldolase/kinase enzyme confers catalytic advantage for C-C bond formation.

A new bifunctional enzyme that displays both aldolase and kinase activities has been designed and successfully used in the synthesis of aldol adducts, employing DHA as initial donor, with an increase in the reaction rate of 20-fold over the parent enzymes, which can be interpreted in terms of substrate channelling.