The Landscape of Osteocalcin Proteoforms Reveals Distinct Structural and Functional Roles of Its Carboxylation Sites.

Human osteocalcin (OC) undergoes reversible, vitamin K-dependent γ-carboxylation at three glutamic acid residues, modulating its release from bones and its hormonal roles. A complete understanding of OC roles and structure-activity relationships is still lacking, as only uncarboxylated and few differently carboxylated variants have been considered so far. To fill this lack of knowledge, a comprehensive experimental and computational investigation of the structural properties and calcium-binding activity of all the OC variants is reported here.

Vibrational spectroscopy coupled with machine learning sheds light on the cellular effects induced by rationally designed TLR4 agonists.

In this work, we present the potential of Fourier transform infrared (FTIR) microspectroscopy to compare on whole cells, in an unbiased and untargeted way, the capacity of bacterial lipopolysaccharide (LPS) and two rationally designed molecules (FP20 and FP20Rha) to activate molecular circuits of innate immunity. These compounds are important drug hits in the development of vaccine adjuvants and tumor immunotherapeutics. The biological assays indicated that FP20Rha was more potent than FP20 in inducing cytokine production in cells and in stimulating IgG antibody production post-vaccination in mice.

Truncation of the constant domain drives amyloid formation by immunoglobulin light chains.

AL amyloidosis is a life-threatening disease caused by deposition of immunoglobulin light chains. While the mechanisms underlying light chains amyloidogenesis in vivo remain unclear, several studies have highlighted the role that tissue environment and structural amyloidogenicity of individual light chains have in the disease pathogenesis. AL natural deposits contain both full-length light chains and fragments encompassing the variable domain (V) as well as different length segments of the constant region (C), thus highlighting the relevance that proteolysis may have in the fibrillogenesis pathway.

Structural insights and aggregation propensity of a super-stable monellin mutant: A new potential building block for protein-based nanostructured materials.

Protein fibrillation is commonly associated with pathologic amyloidosis. However, under appropriate conditions several proteins form fibrillar structures in vitro that can be used for biotechnological applications. MNEI and its variants, firstly designed as single chain derivatives of the sweet protein monellin, are also useful models for protein fibrillary aggregation studies.

Condensation of the N-terminal domain of human topoisomerase 1 is driven by electrostatic interactions and tuned by its charge distribution.

Liquid-liquid phase separation (LLPS) is pivotal in forming biomolecular condensates, which are crucial in several biological processes. Intrinsically disordered regions (IDRs) are typically responsible for driving LLPS due to their multivalency and high content of charged residues that enable the establishment of electrostatic interactions. In our study, we examined the role of charge distribution in the condensation of the disordered N-terminal domain of human topoisomerase I (hNTD).

Modulation of Alpha-Synuclein Conformational Ensemble and Aggregation Pathways by Dopamine and Related Molecules.

Dopaminergic neurons are constantly threatened by the thin boundaries between functional α-synuclein (AS) structural disorder and pathogenic aggregation, and between dopamine (DA) neurotransmitter activity and accumulation of cytotoxic by-products. The possibilities of developing drugs for Parkinson's disease (PD) depend on our understanding of the molecular mechanisms that cause or accompany the pathological structural changes in AS. This review focuses on the three interconnected aspects of AS conformational transitions, its aggregation pathways and ligand binding.

Addressing Critical Issues Related to Storage and Stability of the Vault Nanoparticle Expressed and Purified from .

The vault nanoparticle is a eukaryotic assembly consisting of 78 copies of the 99-kDa major vault protein. They generate two cup-shaped symmetrical halves, which in vivo enclose protein and RNA molecules. Overall, this assembly is mainly involved in pro-survival and cytoprotective functions.

Cheese-whey permeate improves the fitness of Escherichia coli cells during recombinant protein production.

Background: Escherichia coli cells are the most frequently used hosts in recombinant protein production processes and mainly require molecules such as IPTG or pure lactose as inducers of heterologous expression. A possible way to reduce the production costs is to replace traditional inducers with waste materials such as cheese whey permeate (CWP). CWP is a secondary by-product generated from the production of the valuable whey proteins, which are obtained from ultrafiltration of cheese whey, a main by-product of the dairy industry, which is rich in lactose.

An and investigation of cytoplasmic TDP-43 inclusions reveals the absence of a clear amyloid signature.

Several neurodegenerative conditions are associated with a common histopathology within neurons of the central nervous system, consisting of the deposition of cytoplasmic inclusions of TAR DNA-binding protein 43 (TDP-43). Such inclusions have variably been described as morphologically and molecularly ordered aggregates having amyloid properties, as filaments without the cross-β-structure and dye binding specific for amyloid, or as amorphous aggregates with no defined structure and fibrillar morphology. Here we have expressed human full-length TDP-43 in neuroblastoma x spinal cord 34 (NSC-34) cells to investigate the morphological, structural, and tinctorial properties of TDP-43 inclusions .

A Global Picture of Molecular Changes Associated to LPS Treatment in THP-1 Derived Human Macrophages by Fourier Transform Infrared Microspectroscopy.

Macrophages are among the first immune cells involved in the initiation of the inflammatory response to protect the host from pathogens. THP-1 derived macrophages (TDM) are used as a model to study the pro-inflammatory effects of lipopolysaccharide (LPS) exposure. Intact TDM cells were analysed by Fourier transform infrared (FTIR) microspectroscopy, supported by multivariate analysis, to obtain a snapshot of the molecular events sparked by LPS stimulation in macrophage-like cells.

Contribution of Infrared Spectroscopy to the Understanding of Amyloid Protein Aggregation in Complex Systems.

Infrared (IR) spectroscopy is a label-free and non-invasive technique that probes the vibrational modes of molecules, thus providing a structure-specific spectrum. The development of infrared spectroscopic approaches that enable the collection of the IR spectrum from a selected sample area, from micro- to nano-scale lateral resolutions, allowed to extend their application to more complex biological systems, such as intact cells and tissues, thus exerting an enormous attraction in biology and medicine. Here, we will present recent works that illustrate in particular the applications of IR spectroscopy to the characterization of the conformational properties of protein aggregates and to the investigation of the other biomolecules surrounding the amyloids.

Fluorescence of KCl Aqueous Solution: A Possible Spectroscopic Signature of Nucleation.

Ion pairing in water solutions alters both the water hydrogen-bond network and ion solvation, modifying the dynamics and properties of electrolyte water solutions. Here, we report an anomalous intrinsic fluorescence of KCl aqueous solution at room temperature and show that its intensity increases with the salt concentration. From the ab initio density functional theory (DFT) and time-dependent DFT modeling, we propose that the fluorescence emission could originate from the stiffening of the hydrogen bond network in the hydration shell of solvated ion-pairs that suppresses the fast nonradiative decay and allows the slower radiative channel to become a possible decay pathway.

Short-chain alcohols inactivate an immobilized industrial lipase through two different mechanisms.

Broadly used in biocatalysis as acyl acceptors or (co)-solvents, short-chain alcohols often cause irreversible loss of enzyme activity. Understanding the mechanisms of inactivation is a necessary step toward the optimization of biocatalytic reactions and the design of enzyme-based sustainable processes. The functional and structural responses of an immobilized enzyme, Novozym 435 (N-435), exposed to methanol, ethanol, and tert-butanol, are explored in this work.

Characterization of the Conformational Properties of Soluble and Insoluble Proteins by Fourier Transform Infrared Spectroscopy.

The FTIR (micro-)spectroscopy method applied to the study of the structural properties of different soluble and insoluble proteins will be illustrated. In particular, we will discuss the procedure to analyze proteins in form of hydrated films and in solution by means of attenuated total reflection (ATR) measurements. Moreover, we will describe the procedure to characterize bacterial inclusion bodies (IBs) and amyloid deposits within human tissues by means of FTIR microspectroscopy.

Tear-Based Vibrational Spectroscopy Applied to Amyotrophic Lateral Sclerosis.

Biofluid analysis by optical spectroscopy techniques is attracting considerable interest due to its potential to revolutionize diagnostics and precision medicine, particularly for neurodegenerative diseases. However, the lack of effective biomarkers combined with the unaccomplished identification of convenient biofluids has drastically hampered optical advancements in clinical diagnosis and monitoring of neurodegenerative disorders. Here, we show that vibrational spectroscopy applied to human tears opens a new route, offering a non-invasive, label-free identification of a devastating disease such as amyotrophic lateral sclerosis (ALS).

Transcriptomic analysis of Rhodococcus opacus R7 grown on polyethylene by RNA-seq.

Plastic waste management has become a global issue. Polyethylene (PE) is the most abundant synthetic plastic worldwide, and one of the most resistant to biodegradation. Indeed, few bacteria can degrade polyethylene.

Pathological ATX3 Expression Induces Cell Perturbations in as Revealed by Biochemical and Biophysical Investigations.

Amyloid aggregation of human ataxin-3 (ATX3) is responsible for spinocerebellar ataxia type 3, which belongs to the class of polyglutamine neurodegenerative disorders. It is widely accepted that the formation of toxic oligomeric species is primarily involved in the onset of the disease. For this reason, to understand the mechanisms underlying toxicity, we expressed both a physiological (ATX3-Q24) and a pathological ATX3 variant (ATX3-Q55) in a simplified cellular model, .

Conversion of sugar beet residues into lipids by Lipomyces starkeyi for biodiesel production.

Background: Lipids from oleaginous yeasts emerged as a sustainable alternative to vegetable oils and animal fat to produce biodiesel, the biodegradable and environmentally friendly counterpart of petro-diesel fuel. To develop economically viable microbial processes, the use of residual feedstocks as growth and production substrates is required.

Results: In this work we investigated sugar beet pulp (SBP) and molasses, the main residues of sugar beet processing, as sustainable substrates for the growth and lipid accumulation by the oleaginous yeast Lipomyces starkeyi.

Exploring the use of leucine zippers for the generation of a new class of inclusion bodies for pharma and biotechnological applications.

Background: Inclusion bodies (IBs) are biologically active protein aggregates forming natural nanoparticles with a high stability and a slow-release behavior. Because of their nature, IBs have been explored to be used as biocatalysts, in tissue engineering, and also for human and animal therapies. To improve the production and biological efficiency of this nanomaterial, a wide range of aggregation tags have been evaluated.

Conformational Stability and Dynamics in Crystals Recapitulate Protein Behavior in Solution.

A growing body of evidences has established that in many cases proteins may preserve most of their function and flexibility in a crystalline environment, and several techniques are today capable to characterize molecular properties of proteins in tightly packed lattices. Intriguingly, in the case of amyloidogenic precursors, the presence of transiently populated states (hidden to conventional crystallographic studies) can be correlated to the pathological fate of the native fold; the low fold stability of the native state is a hallmark of aggregation propensity. It remains unclear, however, to which extent biophysical properties of proteins such as the presence of transient conformations or protein stability characterized in crystallo reflect the protein behavior that is more commonly studied in solution.

Structural features of the glutamate-binding protein from Corynebacterium glutamicum.

L-glutamate (Glu) is the major excitatory transmitter in mammalian brain. Inadequate concentration of Glu in the brain correlates to mood disorder. In industry, Glu is used as a flavour enhancer in food and in foodstuff processing.