Cloning, Expression and Functional Characterization of CAT2 Arginine Transporter.

The amino acid membrane transporters of grape species take part in metabolic pathways that play crucial roles in nitrogen trafficking and in the synthesis of secondary metabolites. Therefore, identifying these amino acid transporters and defining their functional properties might have further applications in crop improvement and, hence, relevance to human nutrition. The (Cation Amino acid Transporter) transporter cDNA has been isolated and cloned into a specific plasmid for over-expression in .

The antiport mechanism of the human LAT1 transporter: defining the molecular determinants by means of wet and dry approaches.

LAT1 (SLC7A5) catalyzes an antiport of essential amino acids at the Blood Brain Barrier and placenta barrier where it is mainly expressed. LAT1 becomes overexpressed in human cancers, even when originating from tissues in which it is not highly expressed. The antiport mediated by the transporter is crucial for maintaining the essential amino acid homeostasis in the brain, as demonstrated by the occurrence of Autism Spectrum Disorder forms caused by inherited mutations of LAT1.

Impact of 7-ketocholesterol on the function and stability of the LAT1 transporter.

The L-type amino acid transporter (LAT1, SLC7A5) is involved in supplying essential amino acids for brain and fetal development. The regulation of LAT1 function and stability by cholesterol was previously described in ex vivo and in vitro experimental models. The complexity of the relationship between LAT1 and cholesterol has been further studied by investigating the impact of selected oxidized forms of cholesterol, focusing on the most abundant auto-oxidized form of cholesterol, namely 7-ketocholesterol, which increases with age and pathologies characterized by high reactive oxygen species (ROS) levels.

OCTN1 (SLC22A4) as a Target of Heavy Metals: Its Possible Role in Microplastic Threats.

Microplastics represent a threat due to their ability to enter the food chain, with harmful consequences for living organisms. The riskiness of these particles is also linked to the release of other contaminants, such as heavy metals. Solute Carriers (SLCs) represent eminent examples of first-level targets of heavy metals due to their localization on the cell surface.

Lysine 204 is crucial for the antiport function of the human LAT1 transporter.

LAT1 (SLC7A5) catalyzes an antiport reaction of amino acids with specificity towards the essential ones. It is mainly expressed at the Blood Brain Barrier and placenta barriers, but it becomes over-expressed in virtually all human cancers even if originating from tissues with lower expression levels. The antiport reaction of LAT1 is crucial at the BBB since its inherited loss causes Autism Spectrum Disorder.

Carnitine traffic and human fertility.

Carnitine is a vital molecule in human metabolism, prominently involved in fatty acid β-oxidation within mitochondria. Predominantly sourced from dietary intake, carnitine also derives from endogenous synthesis. This review delves into the complex network of carnitine transport and distribution, emphasizing its pivotal role in human fertility.

The Human OCTN Sub-Family: Gene and Protein Structure, Expression, and Regulation.

OCTN1 and OCTN2 are membrane transport proteins encoded by the and genes, respectively. Even though several transcripts have been predicted by bioinformatics for both genes, only one functional protein isoform has been described for each of them. Both proteins are ubiquitous, and depending on the physiopathological state of the cell, their expression is regulated by well-known transcription factors, although some aspects have been neglected.

Structural insights into the bifunctional enzyme human FAD synthase.

Human flavin adenine dinucleotide synthase (hFADS) is a bifunctional, multi-domain enzyme that exhibits both flavin mononucleotide adenylyltransferase and pyrophosphatase activities. Here we report the crystal structure of full-length hFADS2 and its C-terminal PAPS domain in complex with flavin adenine dinucleotide (FAD), and dissect the structural determinants underlying the contribution of each individual domain, within isoforms 1 and 2, to each of the two enzymatic activities. Structural and functional characterization performed on complete or truncated constructs confirmed that the C-terminal domain tightly binds FAD and catalyzes its synthesis, while the combination of the N-terminal molybdopterin-binding and KH domains is the minimal essential substructure required for the hydrolysis of FAD and other ADP-containing dinucleotides.

Inflammation and Organic Cation Transporters Novel (OCTNs).

Inflammation is a physiological condition characterized by a complex interplay between different cells handled by metabolites and specific inflammatory-related molecules. In some pathological situations, inflammation persists underlying and worsening the pathological state. Over the years, two membrane transporters namely OCTN1 (SLC22A4) and OCTN2 (SLC22A5) have been shown to play specific roles in inflammation.

Bacterial over-production of the functionally active human SLC38A2 (SNAT2) exploiting the mistic tag: a cheap and fast tool for testing ligands.

Background: SLC38A2 is a ubiquitously expressed Na-dependent transporter specific for small and medium neutral amino acids. It is involved in human pathologies, such as type II diabetes and cancer. Despite its relevance in human physio-pathology, structure/function relationship studies and identification of ligands with regulatory roles are still in infancy.

Over-Production of the Human SLC7A10 in and Functional Assay in Proteoliposomes.

The human SLC7A10 transporter, also known as ASC-1, catalyzes the transport of some neutral amino acids. It is expressed in astrocytes, neurons, and adipose tissues, playing roles in learning, memory processes, and lipid metabolism, thus being involved in neurological and metabolic pathologies. Structure/function studies on this transporter are still in their infancy.

OCTN1 (SLC22A4) displays two different transport pathways for organic cations or zwitterions.

Background: OCTN1 belongs to the SLC22 family, which includes transporters for cationic, zwitterionic, and anionic substrates. OCTN1 function and role in cells are still poorly understood. Not only cations, such as TEA, but also zwitterions, such as carnitine and ergothioneine, figure among transported molecules.

LAT1 (SLC7A5) catalyzes copper(histidinate) transport switching from antiport to uniport mechanism.

LAT1 (SLC7A5) is one of the most studied membrane transporters due to its relevance to physiology in supplying essential amino acids to brain and fetus, and to pathology being linked to nervous or embryo alterations; moreover, LAT1 over-expression is always associated with cancer development. Thus, LAT1 is exploited as a pro-drug vehicle and as a target for anti-cancer therapy. We here report the identification of a new substrate with pathophysiological implications, i.

Increased demand for FAD synthesis in differentiated and stem pancreatic cancer cells is accomplished by modulating FLAD1 gene expression: the inhibitory effect of Chicago Sky Blue.

FLAD1, along with its FAD synthase (FADS, EC 2.7.7.

TMBIM5 is the Ca /H antiporter of mammalian mitochondria.

Mitochondrial Ca ions are crucial regulators of bioenergetics and cell death pathways. Mitochondrial Ca content and cytosolic Ca homeostasis strictly depend on Ca transporters. In recent decades, the major players responsible for mitochondrial Ca uptake and release have been identified, except the mitochondrial Ca /H exchanger (CHE).

Production of recombinant human xCT (SLC7A11) and reconstitution in proteoliposomes for functional studies.

The plasma membrane transporter xCT belongs to the SLC7 family and has the physiological role of mediating the exchange of glutamate and cystine across the cell plasma membrane, being crucial for redox control. The xCT protein forms a heterodimer with the ancillary protein CD98. Over the years, xCT became a hot pharmacological target due to the documented over-expression in virtually all human cancers, which rely on cystine availability for their progression.

Heterologous (Over) Expression of Human SoLute Carrier (SLC) in Yeast: A Well-Recognized Tool for Human Transporter Function/Structure Studies.

For more than 20 years, yeast has been a widely used system for the expression of human membrane transporters. Among them, more than 400 are members of the largest transporter family, the SLC superfamily. SLCs play critical roles in maintaining cellular homeostasis by transporting nutrients, ions, and waste products.

Bacterial over-expression of functionally active human CT2 (SLC22A16) carnitine transporter.

Background: Escherichia coli is a widely used tool for the over-expression of human proteins for studying structure and function. The toxicity of human proteins for E. coli often hampers the expression.

Strategies for Successful Over-Expression of Human Membrane Transport Systems Using Bacterial Hosts: Future Perspectives.

Ten percent of human genes encode for membrane transport systems, which are key components in maintaining cell homeostasis. They are involved in the transport of nutrients, catabolites, vitamins, and ions, allowing the absorption and distribution of these compounds to the various body regions. In addition, roughly 60% of FDA-approved drugs interact with membrane proteins, among which are transporters, often responsible for pharmacokinetics and side effects.

The Nutraceutical Alliin From Garlic Is a Novel Substrate of the Essential Amino Acid Transporter LAT1 (SLC7A5).

The plasma membrane transporter LAT1 (SLC7A5) is a crucial player for cell homeostasis because it is responsible for providing cells with essential amino acids and hormones. LAT1 forms a functional heterodimer with the cell surface antigen heavy chain CD98 (also known as 4F2hc and SLC3A2), a type II membrane glycoprotein, which is essential for LAT1 stability and localization to the plasma membrane. The relevance of LAT1 for human metabolism is also related to its altered expression in human diseases, such as cancer and diabetes.

OCTN1: A Widely Studied but Still Enigmatic Organic Cation Transporter Linked to Human Pathology and Drug Interactions.

The Novel Organic Cation Transporter, OCTN1, is the first member of the OCTN subfamily; it belongs to the wider Solute Carrier family SLC22, which counts many members including cation and anion organic transporters. The tertiary structure has not been resolved for any cation organic transporter. The functional role of OCNT1 is still not well assessed despite the many functional studies so far conducted.

Chemical Approaches for Studying the Biology and Pharmacology of Membrane Transporters: The Histidine/Large Amino Acid Transporter SLC7A5 as a Benchmark.

The localization of membrane transporters at the forefront of natural barriers makes these proteins very interesting due to their involvement in the absorption and distribution of nutrients and xenobiotics, including drugs. Over the years, structure/function relationship studies have been performed employing several strategies, including chemical modification of exposed amino acid residues. These approaches are very meaningful when applied to membrane transporters, given that these proteins are characterized by both hydrophobic and hydrophilic domains with a different degree of accessibility to employed chemicals.

ASCT1 and ASCT2: Brother and Sister?

The SLC1 family includes seven members divided into two groups, namely, EAATs and ASCTs, that share similar 3D architecture; the first one includes high-affinity glutamate transporters, and the second one includes SLC1A4 and SLC1A5, known as ASCT1 and ASCT2, respectively, responsible for the traffic of neutral amino acids across the cell plasma membrane. The physiological role of ASCT1 and ASCT2 has been investigated over the years, revealing different properties in terms of substrate specificities, affinities, and regulation by physiological effectors and posttranslational modifications. Furthermore, ASCT1 and ASCT2 are involved in pathological conditions, such as neurodegenerative disorders and cancer.

Heterologous Overexpression of Human FAD Synthase Isoforms 1 and 2.

The study of human FAD synthase enzymes requires a recombinant strategy to produce large amount of purified proteins in a soluble form. E. coli was exploited to this aim.

Carnitine Traffic in Cells. Link With Cancer.

Metabolic flexibility is a peculiar hallmark of cancer cells. A growing number of observations reveal that tumors can utilize a wide range of substrates to sustain cell survival and proliferation. The diversity of carbon sources is indicative of metabolic heterogeneity not only across different types of cancer but also within those sharing a common origin.