Unraveling the mechanism of glutamate transport

Glutamate concentration in the synaptic cleft is tightly regulated by a family of secondary active transporters called excitatory amino acid transporters (EAAT1-5) that are able to symport three Na+ ions and one H+ and counter-transport one K+ ion, with uncoupled chloride conductance. In this thesis, the studies have been conducted on the archeal homologue, GltTk.During the initial steps of binding, the Na+ ions entering the binding pocket from the bulk most likely follow a series of transient bindings with other amino acid residues before reaching their final states. This was revealed by combination of atomistic-scale MD simulations including the application of Markov state model and functional studies. I present the attempts of creating larger nanodiscs for structural purposes using three different approaches: two by extracting the protein directly from the membranes with the diisobutylene/maleic acid copolymer, called DIBMA, and one by changing the reconstitution ratios of the protein with the protein belt MSP2N2 and the lipids. During the transport cycle, an aqueous channel between the scaffold and the transport domain forms and it allows the thermodynamically uncoupled anionic flux. This state is called the chloride-conductive state (ClCS). I present the characterization of the chloride channel in GltTk using solid-supported membrane electrophysiology (SSM). This same electrophysiological method was used to study the implications of the single point mutation of the only proline present in TM 5 to arginine, that in humans is responsible for the neurological disorder called episodic ataxia 6

Cysteine-mediated decyanation of vitamin B12 by the predicted membrane transporter BtuM

Uptake of vitamin B12 is essential for many prokaryotes, but in most cases the membrane proteins involved are yet to be identified. We present the biochemical characterization and high-resolution crystal structure of BtuM, a predicted bacterial vitamin B12 uptake system. BtuM binds vitamin B12 in its base-off conformation, with a cysteine residue as axial ligand of the corrin cobalt ion. Spectroscopic analysis indicates that the unusual thiolate coordination allows for decyanation of vitamin B12. Chemical modification of the substrate is a property other characterized vitamin B12-transport proteins do not exhibit

Acute Delta Hepatitis in Italy spanning three decades (1991–2019): Evidence for the effectiveness of the hepatitis B vaccination campaign

Updated incidence data of acute Delta virus hepatitis (HDV) are lacking worldwide. Our aim was to evaluate incidence of and risk factors for acute HDV in Italy after the introduction of the compulsory vaccination against hepatitis B virus (HBV) in 1991. Data were obtained from the National Surveillance System of acute viral hepatitis (SEIEVA). Independent predictors of HDV were assessed by logistic-regression analysis. The incidence of acute HDV per 1-million population declined from 3.2 cases in 1987 to 0.04 in 2019, parallel to that of acute HBV per 100,000 from 10.0 to 0.39 cases during the same period. The median age of cases increased from 27 years in the decade 1991-1999 to 44 years in the decade 2010-2019 (p < .001). Over the same period, the male/female ratio decreased from 3.8 to 2.1, the proportion of coinfections increased from 55% to 75% (p = .003) and that of HBsAg positive acute hepatitis tested for by IgM anti-HDV linearly decreased from 50.1% to 34.1% (p < .001). People born abroad accounted for 24.6% of cases in 2004-2010 and 32.1% in 2011-2019. In the period 2010-2019, risky sexual behaviour (O.R. 4.2; 95%CI: 1.4-12.8) was the sole independent predictor of acute HDV; conversely intravenous drug use was no longer associated (O.R. 1.25; 95%CI: 0.15-10.22) with this. In conclusion, HBV vaccination was an effective measure to control acute HDV. Intravenous drug use is no longer an efficient mode of HDV spread. Testing for IgM-anti HDV is a grey area requiring alert. Acute HDV in foreigners should be monitored in the years to come

Rinascimento: crescere perseguendo un sogno

Non present

Scout: un percorso tra istinto e ricerca

Non present

Mutation in glutamate transporter homologue GltTk provides insights into pathologic mechanism of episodic ataxia 6

Episodic ataxias (EAs) are rare neurological conditions affecting the nervous system and typically leading to motor impairment. EA6 is linked to the mutation of a highly conserved proline into an arginine in the glutamate transporter EAAT1. In vitro studies showed that this mutation leads to a reduction in the substrates transport and an increase in the anion conductance. It was hypothesised that the structural basis of these opposed functional effects might be the straightening of transmembrane helix 5, which is kinked in the wild-type protein. In this study, we present the functional and structural implications of the mutation P208R in the archaeal homologue of glutamate transporters GltTk. We show that also in GltTk the P208R mutation leads to reduced aspartate transport activity and increased anion conductance, however a cryo-EM structure reveals that the kink is preserved. The arginine side chain of the mutant points towards the lipidic environment, where it may engage in interactions with the phospholipids, thereby potentially interfering with the transport cycle and contributing to stabilisation of an anion conducting state

Enteric Alpha-Synuclein Inclusions, Colonic Inflammation, Increased Mucosal Permeability and Alterations of Bowel Neuromuscular Functions Precede Central Neurodegeneration in a Transgenic Mouse Model of Parkinson's Disease

No abstract availabl