Expanding SPG18 clinical spectrum: autosomal dominant mutation causes complicated hereditary spastic paraplegia in a large family.

Background: SPG18 is caused by mutations in the endoplasmic reticulum lipid raft associated 2 (ERLIN2) gene. Autosomal recessive (AR) mutations are usually associated with complicated hereditary spastic paraplegia (HSP), while autosomal dominant (AD) mutations use to cause pure SPG18.

Aim: To define the variegate clinical spectrum of the SPG18 and to evaluate a dominant negative effect of erlin2 (encoded by ERLIN2) on oligomerization as causing differences between AR and AD phenotypes.

Molecular Resonance Imaging of the CAIX Expression in Mouse Mammary Adenocarcinoma Cells.

The carbonic anhydrase isoform IX (hCAIX) is one of the main players in extracellular tumor pH regulation, and it is known to be overexpressed in breast cancer and other common tumors. hCA IX supports the growth and survival of tumor cells, and its expression is correlated with metastasis and resistance to therapies, making it an interesting biomarker for diagnosis and therapy. The aim of this work deals with the development of an MRI imaging probe able to target the extracellular non-catalytic proteoglycan-like (PG) domain of CAIX.

Bone Marrow Mesenchymal Stromal/Stem Cell-Derived Extracellular Vesicles Promote Corneal Wound Repair by Regulating Inflammation and Angiogenesis.

Severe corneal damage leads to complete vision loss, thereby affecting life quality and impinging heavily on the healthcare system. Current clinical approaches to manage corneal wounds suffer from severe drawbacks, thus requiring the development of alternative strategies. Of late, mesenchymal stromal/stem cell (MSC)-derived extracellular vesicles (EVs) have become a promising tool in the ophthalmic field.

Regenerative Approaches and Future Trends for the Treatment of Corneal Burn Injuries.

Ocular chemical and thermal burns are frequent causes of hospitalization and require immediate interventions and care. Various surgical and pharmacological treatment strategies are employed according to damage severity. Controlling inflammation and neovascularization while promoting normal ocular surface anatomy and function restoration is the principal aim.

A Late-Stage Synthetic Approach to Lanthionine-Containing Peptides via S-Alkylation on Cyclic Sulfamidates Promoted by Molecular Sieves.

A one-pot, high-yield procedure for synthesizing lanthionine-containing peptides was developed. It relies on the S-alkylation of cysteine-containing peptides with chiral cyclic sulfamidates. The key feature of this approach is the use of mild reaction conditions (only activated molecular sieves are employed as the catalyst), leading to good chemoselectivity and excellent stereochemical control.

Chemoselective Glycosylation of Peptides through S-Alkylation Reaction.

An efficient and rapid procedure for synthesizing S-linked glycopeptides is reported. The approach uses activated molecular sieves as a base to promote the selective S-alkylation of readily prepared cysteine-containing peptides, upon reaction of appropriate glycosyl halides. Considering the very mild conditions employed, the chemoselective linkage of the electrophilic sugar with a peptide sulfhydryl group occurred in satisfactory yield, allowing the incorporation of mono and disaccharide moieties.

TRPV4 mutations in children with congenital distal spinal muscular atrophy.

Inherited disorders characterized by motor neuron loss and muscle weakness are genetically heterogeneous. The recent identification of mutations in the gene encoding transient receptor potential vanilloid 4 (TRPV4) in distal spinal muscular atrophy (dSMA) prompted us to screen for TRPV4 mutations in a small group of children with compatible phenotype. In a girl with dSMA and vocal cord paralysis, we detected a new variant (p.

Novel Gd(III)-based probes for MR molecular imaging of matrix metalloproteinases.

Two novel Gd-based contrast agents (CAs) for the molecular imaging of matrix metalloproteinases (MMPs) were synthetized and characterized in vitro and in vivo. These probes were based on the PLG*LWAR peptide sequence, known to be hydrolyzed between Gly and Leu by a broad panel of MMPs. A Gd-DOTA chelate was conjugated to the N-terminal position through an amide bond, either directly to proline (compd Gd-K11) or through a hydrophilic spacer (compd Gd-K11N).

In vivo labeling of B16 melanoma tumor xenograft with a thiol-reactive gadolinium based MRI contrast agent.

Murine melanoma B16 cells display on the extracellular side of the plasma membrane a large number of reactive protein thiols (exofacial protein thiols, EPTs). These EPTs can be chemically labeled with Gd-DO3A-PDP, a Gd(III)-based MRI contrast agent bearing a 2-pyridinedithio chemical function for the recognition of EPTs. Uptake of gadolinium up to 10(9) Gd atoms per cell can be achieved.

Gadolinium-doped LipoCEST agents: a potential novel class of dual 1H-MRI probes.

A novel class of paramagnetic liposome-based systems acting as dual T(1) and CEST (1)H-MRI contrast agents is described. The vesicles contain a shift reagent in the aqueous core and a Gd-complex on the external surface conjugated through a biodegradable linker. As such, the probe can generate T(1) contrast only, but after the cleavage and removal of the Gd-coating, the CEST contrast is switched on.

Exofacial protein thiols as a route for the internalization of Gd(III)-based complexes for magnetic resonance imaging cell labeling.

Four novel MRI Gd(III)-based probes have been synthesized and evaluated for their labeling properties on cultured cell lines K562, C6, and B16. The labeling strategy relies upon the fact that cells display a large number of reactive exofacial protein thiols (EPTs) that can be exploited as anchorage points for suitably activated MRI probes. The probes are composed of a Gd(III) chelate (based on either DO3A or DTPA) connected through a flexible linker to the 2-pyridyldithio chemical function for binding to EPTs.

Targeting exofacial protein thiols with Gd(III) complexes. An efficient procedure for MRI cell labelling.

Cells display on the outer surface of the plasma membrane a large number of protein thiols that can be reversibly labelled with suitably designed Gd(III)-based contrast agents for cell tracking by MRI.

Structural and functional differences between KRIT1A and KRIT1B isoforms: a framework for understanding CCM pathogenesis.

KRIT1 is a disease gene responsible for Cerebral Cavernous Malformations (CCM). It encodes for a protein containing distinct protein-protein interaction domains, including three NPXY/F motifs and a FERM domain. Previously, we isolated KRIT1B, an isoform characterized by the alternative splicing of the 15th coding exon and suspected to cause CCM when abnormally expressed.

Functional and structural features of the oxyanion hole in a thermophilic esterase from Alicyclobacillus acidocaldarius.

Recent mutagenic and molecular modelling studies suggested a role for glycine 84 in the putative oxyanion loop of the carboxylesterase EST2 from Alicyclobacillus acidocaldarius. A 114 times decrease of the esterase catalytic activity of the G84S mutant was observed, without changes in the thermal stability. The recently solved three-dimensional (3D) structure of EST2 in complex with a HEPES molecule permitted to demonstrate that G84 (together with G83 and A156) is involved in the stabilization of the oxyanion through a hydrogen bond from its main chain NH group.

Carbonic anhydrase inhibitors: X-ray crystallographic studies for the binding of 5-amino-1,3,4-thiadiazole-2-sulfonamide and 5-(4-amino-3-chloro-5-fluorophenylsulfonamido)-1,3,4-thiadiazole-2-sulfonamide to human isoform II.

The X-ray crystal structures of 5-amino-1,3,4-thiadiazole-2-sulfonamide (the acetazolamide precursor) and 5-(4-amino-3-chloro-5-fluorophenylsulfonamido)-1,3,4-thiadiazole-2-sulfonamide in complex with the human isozyme II of carbonic anhydrase (CA, EC 4.2.1.

Carbonic anhydrase inhibitors: stacking with Phe131 determines active site binding region of inhibitors as exemplified by the X-ray crystal structure of a membrane-impermeant antitumor sulfonamide complexed with isozyme II.

Structure for the adduct of carbonic anhydrase II with 1-N-(4-sulfamoylphenyl-ethyl)-2,4,6-trimethylpyridinium perchlorate, a membrane-impermeant antitumor sulfonamide, is reported. The phenylethyl moiety fills the active site, making van der Waals interactions with side chains of Gln192, Val121, Phe131, Leu198, Thr200. The 2,4,6-trimethylpyridinium functionality is at van der Waals distance from the aliphatic chain of Ile91 being involved in strong offset face-to-face stacking with Phe131.

Carbonic anhydrase inhibitors. Zonisamide is an effective inhibitor of the cytosolic isozyme II and mitochondrial isozyme V: solution and X-ray crystallographic studies.

The antiepileptic drug zonisamide was considered to act as a weak inhibitor of the zinc enzyme carbonic anhydrase (CA, EC 4.2.1.

Carbonic anhydrase inhibitors: X-ray crystal structure of a benzenesulfonamide strong CA II and CA IX inhibitor bearing a pentafluorophenylaminothioureido tail in complex with isozyme II.

N-1-(4-Sulfamoylphenyl)-N-4-pentafluorophenyl-thiosemicarbazide was prepared by the reaction of 4-isothiocyanato-benzenesulfonamide with pentafluorophenyl hydrazine, and proved to be an effective inhibitor of several isozymes of the zinc enzyme carbonic anhydrase (CA, EC 4.2.1.

The crystal structure of an EST2 mutant unveils structural insights on the H group of the carboxylesterase/lipase family.

Esterase 2 (EST2) from the thermophilic eubacterium Alicyclobacillus acidocaldarius is a thermostable serine hydrolase belonging to the H group of the esterase/lipase family. This enzyme hydrolyzes monoacylesters of different acyl-chain length and various compounds with industrial interest. EST2 displays an optimal temperature at 70 degrees C and maximal activity with pNP-esters having acyl-chain bearing from six to eight carbon atoms.

A substrate-induced switch in the reaction mechanism of a thermophilic esterase: kinetic evidences and structural basis.

The reaction mechanism of the esterase 2 (EST2) from Alicyclobacillus acidocaldarius was studied at the kinetic and structural level to shed light on the mechanism of activity and substrate specificity increase previously observed in its double mutant M211S/R215L. In particular, the values of kinetic constants (k1, k(-1), k2, and k3) along with activation energies (E1, E(-1), E2, and E3) were measured for wild type and mutant enzyme. The previously suggested substrate-induced switch in the reaction mechanism from kcat=k3 with a short acyl chain substrate (p-nitrophenyl hexanoate) to kcat=k2 with a long acyl chain substrate (p-nitrophenyl dodecanoate) was validated.

Insights into peptide nucleic acid (PNA) structural features: the crystal structure of a D-lysine-based chiral PNA-DNA duplex.

Peptide nucleic acids (PNAs) are oligonucleotide analogues in which the sugar-phosphate backbone has been replaced by a pseudopeptide skeleton. They bind DNA and RNA with high specificity and selectivity, leading to PNA-RNA and PNA-DNA hybrids more stable than the corresponding nucleic acid complexes. The binding affinity and selectivity of PNAs for nucleic acids can be modified by the introduction of stereogenic centers (such as D-Lys-based units) into the PNA backbone.

Crystallization and preliminary X-ray diffraction studies of Aes acetyl-esterase from Escherichia coli.

The acetyl-esterase Aes from Escherichia coli, which belongs to the HSL group of the esterase/lipase superfamily, has been crystallized by the hanging-drop vapour-diffusion method using polyethylene glycol 8000 as a precipitant and magnesium chloride as an additive. Crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 110.0, b = 190.