Disentangling performance-monitoring signals encoded in feedback-related EEG dynamics.

The feedback-related negativity (FRN) is a well-established electrophysiological correlate of feedback-processing. However, there is still an ongoing debate whether the FRN is driven by negative or positive reward prediction errors (RPE), valence of feedback, or mere surprise. Our study disentangles independent contributions of valence, surprise, and RPE on the feedback-related neuronal signal including the FRN and P3 components using the statistical power of a sample of N = 992 healthy individuals. The participants performed a modified time-estimation task, while EEG from 64 scalp electrodes was recorded. Our results show that valence coding is present during the FRN with larger amplitudes for negative feedback. The FRN is further modulated by surprise in a valence-dependent way being more positive-going for surprising positive outcomes. The P3 was strongly driven by both global and local surprise, with larger amplitudes for unexpected feedback and local deviants. Behavioral adaptations after feedback and FRN just show small associations. Results support the theory of the FRN as a representation of a signed RPE. Additionally, our data indicates that surprising positive feedback enhances the EEG response in the time window of the P3. These results corroborate previous findings linking the P3 to the evaluation of PEs in decision making and learning tasks.

Laser Structuring and DLC Coating of Elastomers for High Performance Applications.

Even though hard, low friction coatings such as diamond like carbon (DLC) would be beneficial for the performance and longevity of rubber seals, a crucial challenge remains. The elastic mismatch of rubber substrate and DLC coating prevents a fracture free coating application. In this work, a nature inspired approach is applied to render the stiff coating flexible and resilient to delamination at the same time by direct patterning. Rubber substrates were laser structured with tile patterns and subsequently DLC coated. Tensile and tribology tests were performed on structured and unstructured samples. Unstructured DLC coatings showed a crack pattern induced by the coating process, which was further fragmented by tensile stress. Coatings with tile patterns did not experience a further fragmentation under load. During continuous tribological loading, less heterogenous damage is produced for tile structured samples. The findings are ascribed to the relief of induced coating stress by the tile structure, meaning a more resilient coating.

Anodal Transcranial Direct Current Stimulation Over S1 Differentially Modulates Proprioceptive Accuracy in Young and Old Adults.

BACKGROUND: Proprioception is a prerequisite for successful motor control but declines throughout the lifespan. Brain stimulation techniques such as anodal transcranial direct current stimulation (a-tDCS) are capable of enhancing sensorimotor performance across different tasks and age groups. Despite such growing evidence for a restorative potential of tDCS, its impact on proprioceptive accuracy has not been studied in detail yet. OBJECTIVE: This study investigated online effects of a-tDCS over S1 on proprioceptive accuracy in young (YA) and old healthy adults (OA). METHODS: The effect of 15 min of a-tDCS vs. sham on proprioceptive accuracy was assessed in a cross-over, double blind experiment in both age groups. Performance changes were tested using an arm position matching task in a robotic environment. Electrical field (EF) strengths in the target area S1 and control areas were assessed based on individualized simulations. RESULTS: a-tDCS elicited differential changes in proprioceptive accuracy and EF strengths in the two groups: while YA showed a slight improvement, OA exhibited a decrease in performance during a-tDCS. Stronger EF were induced in target S1 and control areas in the YA group. However, no relationship between EF strength and performance change was found. CONCLUSION: a-tDCS over S1 elicits opposing effects on proprioceptive accuracy as a function of age, a result that is important for future studies investigating the restorative potential of a-tDCS in healthy aging and in the rehabilitation of neurological diseases that occur at advanced age. Modeling approaches could help elucidate the relationship between tDCS protocols, brain structure and performance modulation.

Complex Stability During the Transport Cycle of a Subclass I ECF Transporter.

The mechanism of energy-coupling factor (ECF) transporters, a special type of ATP-binding-cassette importers for micronutrients in prokaryotes, is a matter of controversial discussion. Among subclass II ECF transporters, a single ECF interacts with several substrate-binding integral membrane proteins (S units) for individual solutes. Release and catch of the S unit, previously observed experimentally for a subclass II system, was proposed as the mechanism of all ECF transporters. The BioM2NY biotin transporter is a prototype of subclass I systems, among which the S unit is dedicated to a specific ECF. Here we simulated the transport cycle using purified BioM2NY in detergent solution. BioM2NY complexes were stable during all steps. ATP binding was a prerequisite for biotin capture and ATP hydrolysis for subsequent biotin release. The data demonstrate that S units of subclass I ECF transporters do not have to dissociate from holotransporter complexes for high-affinity substrate binding, indicating mechanistic differences between the two subclasses.

ATP-dependent Conformational Changes Trigger Substrate Capture and Release by an ECF-type Biotin Transporter.

Energy-coupling factor (ECF) transporters for vitamins and metal ions in prokaryotes consist of two ATP-binding cassette-type ATPases, a substrate-specific transmembrane protein (S component) and a transmembrane protein (T component) that physically interacts with the ATPases and the S component. The mechanism of ECF transporters was analyzed upon reconstitution of a bacterial biotin transporter into phospholipid bilayer nanodiscs. ATPase activity was not stimulated by biotin and was only moderately reduced by vanadate. A non-hydrolyzable ATP analog was a competitive inhibitor. As evidenced by cross-linking of monocysteine variants and by site-specific spin labeling of the Q-helix followed by EPR-based interspin distance analyses, closure and reopening of the ATPase dimer (BioM2) was a consequence of ATP binding and hydrolysis, respectively. A previously suggested role of a stretch of small hydrophobic amino acid residues within the first transmembrane segment of the S units for S unit/T unit interactions was structurally and functionally confirmed for the biotin transporter. Cross-linking of this segment in BioY (S) using homobifunctional thiol-reactive reagents to a coupling helix of BioN (T) indicated a reorientation rather than a disruption of the BioY/BioN interface during catalysis. Fluorescence emission of BioY labeled with an environmentally sensitive fluorophore was compatible with an ATP-induced reorientation and consistent with a hypothesized toppling mechanism. As demonstrated by [(3)H]biotin capture assays, ATP binding stimulated substrate capture by the transporter, and subsequent ATP hydrolysis led to substrate release. Our study represents the first experimental insight into the individual steps during the catalytic cycle of an ECF transporter in a lipid environment.

Transport of nickel and cobalt ions into bacterial cells by S components of ECF transporters.

Energy-coupling factor (ECF) transporters form a distinct group of ABC-type micronutrient importers in prokaryotes that do not contain extracytoplasmic, soluble substrate-binding proteins. Instead, they consist of a transmembrane substrate-specific S component that interacts with a module composed of a moderately conserved transmembrane (T) component and ABC ATPases. The majority of S components is considered to act as high-affinity binding proteins that strictly depend on their cognate T and ATPase units for transport activity. For a fraction of biotin-specific S units, however, transport activity was demonstrated in their solitary state. Here, we compared the activities of nickel- and cobalt-specific ECF transporters in the presence and absence of their T and ATPase units. Accumulation assays with radioactive metal ions showed that the truncated transporters led to approx. 25% of cell-bound radioactivity compared to the holotransporters. Activity of urease, an intracellular nickel-dependent enzyme, was used as a reporter and clearly indicated that the cell-bound radioactivity correlates with the cytoplasmic metal concentration. The results demonstrate that S units of metal transporters not only bind their substrates on the cell surface but mediate transport across the membrane, a finding of general importance on the way to understand the mechanism of ECF transporters.

A versatile Escherichia coli strain for identification of biotin transporters and for biotin quantification.

Biotin is an essential cofactor of carboxylase enzymes in all kingdoms of life. The vitamin is produced by many prokaryotes, certain fungi, and plants. Animals depend on biotin uptake from their diet and in humans lack of the vitamin is associated with serious disorders. Many aspects of biotin metabolism, uptake, and intracellular transport remain to be elucidated. In order to characterize the activity of novel biotin transporters by a sensitive assay, an Escherichia coli strain lacking both biotin synthesis and its endogenous high-affinity biotin importer was constructed. This strain requires artificially high biotin concentrations for growth. When only trace levels of biotin are available, it is viable only if equipped with a heterologous high-affinity biotin transporter. This feature was used to ascribe transport activity to members of the BioY protein family in previous work. Here we show that this strain together with its parent is also useful as a diagnostic tool for wide-concentration-range bioassays.

Planar substrate-binding site dictates the specificity of ECF-type nickel/cobalt transporters.

The energy-coupling factor (ECF) transporters are multi-subunit protein complexes that mediate uptake of transition-metal ions and vitamins in about 50% of the prokaryotes, including bacteria and archaea. Biological and structural studies have been focused on ECF transporters for vitamins, but the molecular mechanism by which ECF systems transport metal ions from the environment remains unknown. Here we report the first crystal structure of a NikM, TtNikM2, the substrate-binding component (S component) of an ECF-type nickel transporter from Thermoanaerobacter tengcongensis. In contrast to the structures of the vitamin-specific S proteins with six transmembrane segments (TSs), TtNikM2 possesses an additional TS at its N-terminal region, resulting in an extracellular N-terminus. The highly conserved N-terminal loop inserts into the center of TtNikM2 and occludes a region corresponding to the substrate-binding sites of the vitamin-specific S components. Nickel binds to NikM via its coordination to four nitrogen atoms, which are derived from Met1, His2 and His67 residues. These nitrogen atoms form an approximately square-planar geometry, similar to that of the metal ion-binding sites in the amino-terminal Cu(2+)- and Ni(2+)-binding (ATCUN) motif. Replacements of residues in NikM contributing to nickel coordination compromised the Ni-transport activity. Furthermore, systematic quantum chemical investigation indicated that this geometry enables NikM to also selectively recognize Co(2+). Indeed, the structure of TtNikM2 containing a bound Co(2+) ion has almost no conformational change compared to the structure that contains a nickel ion. Together, our data reveal an evolutionarily conserved mechanism underlying the metal selectivity of EcfS proteins, and provide insights into the ion-translocation process mediated by ECF transporters.

Solitary BioY proteins mediate biotin transport into recombinant Escherichia coli.

Energy-coupling factor (ECF) transporters form a large group of vitamin uptake systems in prokaryotes. They are composed of highly diverse, substrate-specific, transmembrane proteins (S units), a ubiquitous transmembrane protein (T unit), and homo- or hetero-oligomeric ABC ATPases. Biotin transporters represent a special case of ECF-type systems. The majority of the biotin-specific S units (BioY) is known or predicted to interact with T units and ABC ATPases. About one-third of BioY proteins, however, are encoded in organisms lacking any recognizable T unit. This finding raises the question of whether these BioYs function as transporters in a solitary state, a feature ascribed to certain BioYs in the past. To address this question in living cells, an Escherichia coli K-12 derivative deficient in biotin synthesis and devoid of its endogenous high-affinity biotin transporter was constructed as a reference strain. This organism is particularly suited for this purpose because components of ECF transporters do not naturally occur in E. coli K-12. The double mutant was viable in media containing either high levels of biotin or a precursor of the downstream biosynthetic path. Importantly, it was nonviable on trace levels of biotin. Eight solitary bioY genes of proteobacterial origin were individually expressed in the reference strain. Each of the BioYs conferred biotin uptake activity on the recombinants, which was inferred from uptake assays with [(3)H]biotin and growth of the cells on trace levels of biotin. The results underscore that solitary BioY transports biotin across the cytoplasmic membrane.

Essential amino acid residues of BioY reveal that dimers are the functional S unit of the Rhodobacter capsulatus biotin transporter.

Energy-coupling factor transporters are a large group of importers for trace nutrients in prokaryotes. The in vivo oligomeric state of their substrate-specific transmembrane proteins (S units) is a matter of debate. Here we focus on the S unit BioY of Rhodobacter capsulatus, which functions as a low-affinity biotin transporter in its solitary state. To analyze whether oligomerization is a requirement for function, a tail-to-head-linked BioY dimer was constructed. Monomeric and dimeric BioY conferred comparable biotin uptake activities on recombinant Escherichia coli. Fluorophore-tagged variants of the dimer were shown by fluorescence anisotropy analysis to oligomerize in vivo. Quantitative mass spectrometry identified biotin in the purified proteins at a stoichiometry of 1:2 for the BioY monomer and 1:4 (referring to single BioY domains) for the dimer. Replacement of the conserved Asp164 (by Asn) and Lys167 (by Arg or Gln) in the monomer and in both halves of the dimer inactivated the proteins. The presence of those mutations in one half of the dimers only slightly affected biotin binding but reduced transport activity to 25% (Asp164Asn and Lys167Arg) or 75% (Lys167Gln). Our data (i) suggest that intermolecular interactions of domains from different dimers provide functionality, (ii) confirm an oligomeric architecture of BioY in living cells, and (iii) demonstrate an essential role of the last transmembrane helix in biotin recognition.