Exploration of the spontaneous fluctuating activity of single enzyme molecules

Single enzyme molecules display inevitable, stochastic fluctuations in their catalytic activity. In metabolism, for instance, the stochastic activity of individual enzymes is averaged out due to their high copy numbers per single cell. However, many processes inside cells rely on single enzyme activity, such as transcription, replication, translation, and histone modifications. Here we introduce the main theoretical concepts of stochastic single-enzyme activity starting from the Michaelis–Menten enzyme mechanism. Next, we discuss stochasticity of multi-substrate enzymes, of enzymes and receptors with multiple conformational states and finally, how fluctuations in receptor activity arise from fluctuations in signal concentration. This paper aims to introduce the exciting field of single-molecule enzyme kinetics and stochasticity to a wider audience of biochemists and systems biologists

Microtubule depolymerization by the kinesin-8 motor Kip3p: a mathematical model

Proteins from the kinesin-8 family promote microtubule (MT) depolymerization, a process thought to be important for the control of microtubule length in living cells. In addition to this MT shortening activity, kinesin 8s are motors that show plus-end directed motility on MTs. Here we describe a simple model that incorporates directional motion and destabilization of the MT plus end by kinesin 8. Our model quantitatively reproduces the key features of length-vs-time traces for stabilized MTs in the presence of purified kinesin 8, including length-dependent depolymerization. Comparison of model predictions with experiments suggests that kinesin 8 depolymerizes processively, i.e., one motor can remove multiple tubulin dimers from a stabilized MT. Fluctuations in MT length as a function of time are related to depolymerization processivity. We have also determined the parameter regime in which the rate of MT depolymerization is length dependent: length-dependent depolymerization occurs only when MTs are sufficiently short; this crossover is sensitive to the bulk motor concentration.Comment: 34 pages, 11 figure

Contrasting Phenotypes in Resistance to Thyroid Hormone Alpha Correlate with Divergent Properties of Thyroid Hormone Receptor α1 Mutant Proteins.

BACKGROUND: Resistance to thyroid hormone alpha (RTHα), a disorder characterized by tissue-selective hypothyroidism and near-normal thyroid function tests due to thyroid receptor alpha gene mutations, is rare but probably under-recognized. This study sought to correlate the clinical characteristics and response to thyroxine (T4) therapy in two adolescent RTHα patients with the properties of the THRA mutation, affecting both TRα1 and TRα2 proteins, they harbored. METHODS: Clinical, auxological, biochemical, and physiological parameters were assessed in each patient at baseline and after T4 therapy. RESULTS: Heterozygous THRA mutations occurring de novo were identified in a 17-year-old male (patient P1; c.788C>T, p.A263V mutation) investigated for mild pubertal delay and in a 15-year-old male (patient P2; c.821T>C, p.L274P mutation) with short stature (0.4th centile), skeletal dysplasia, dysmorphic facies, and global developmental delay. Both individuals exhibited macrocephaly, delayed dentition, and constipation, together with a subnormal T4/triiodothyronine (T3) ratio, low reverse T3 levels, and mild anemia. When studied in vitro, A263V mutant TRα1 was transcriptionally impaired and inhibited the function of its wild-type counterpart at low (0.01-10 nM) T3 levels, with higher T3 concentrations (100 nM-1 μM) reversing dysfunction and such dominant negative inhibition. In contrast, L274P mutant TRα1 was transcriptionally inert, exerting significant dominant negative activity, only overcome with 10 μM of T3. Mirroring this, normal expression of KLF9, a TH-responsive target gene, was achieved in A263V mutation-containing peripheral blood mononuclear cells following 1 μM of T3 exposure, but with markedly reduced expression levels in L274P mutation-containing peripheral blood mononuclear cells, even with 10 μM of T3. Following T4 therapy, growth, body composition, dyspraxia, and constipation improved in P1, whereas growth retardation and constipation in P2 were unchanged. Neither A263V nor L274P mutations exhibited gain or loss of function in the TRα2 background, and no additional phenotype attributable to this was discerned. CONCLUSIONS: This study correlates a milder clinical phenotype and favorable response to T4 therapy in a RTHα patient (P1) with heterozygosity for mutant TRα1 exhibiting partial, T3-reversible, loss of function. In contrast, a more severe clinical phenotype refractory to hormone therapy was evident in another case (P2) associated with severe, virtually irreversible, dysfunction of mutant TRα1

Hyperthyroxinemia and Hypercortisolemia due to Familial Dysalbuminemia.

A 23-year-old man and his grandmother with hyperthyroxinemia and hypercortisolemia were heterozygous for an ALB mutation (p. Arg218Pro), known to cause familial dysalbuminemic hyperthyroxinemia (FDH). However, serum-free cortisol levels in these individuals were normal and total cortisol concentrations fell markedly after depletion of albumin from their serum. We conclude that binding of steroid as well as iodothyronines to mutant albumin causes raised circulating cortisol as well as thyroid hormones in euthyroid euadrenal individuals with R218P FDH, with potential for misdiagnosis, unnecessary investigation, and inappropriate treatment.Wellcome Trust NIHR Cambridge Biomedical Research Centr

How Psychological Stress Affects Emotional Prosody

We explored how experimentally induced psychological stress affects the production and recognition of vocal emotions. In Study 1a, we demonstrate that sentences spoken by stressed speakers are judged by naive listeners as sounding more stressed than sentences uttered by non-stressed speakers. In Study 1b, negative emotions produced by stressed speakers are generally less well recognized than the same emotions produced by non-stressed speakers. Multiple mediation analyses suggest this poorer recognition of negative stimuli was due to a mismatch between the variation of volume voiced by speakers and the range of volume expected by listeners. Together, this suggests that the stress level of the speaker affects judgments made by the receiver. In Study 2, we demonstrate that participants who were induced with a feeling of stress before carrying out an emotional prosody recognition task performed worse than non-stressed participants. Overall, findings suggest detrimental effects of induced stress on interpersonal sensitivity

Detection of Bioactive Compounds in the Mucus Nets of Dendropoma maxima, Sowerby 1825 (Prosobranch Gastropod Vermetidae, Mollusca)

The sessile suspension-feeding wormsnail Dendropoma maxima, Sowerby 1825 (Vermetidae) secretes a mucus net to capture planktonic prey. The nets are spread out over the corals and often have remarkable deleterious effects on them like changes in growth form and pigmentation shifts not uncommonly resulting in tissue necrosis. Until now, there is no explanation for this phenomenon although the indication as well as theories about its genesis is mentioned in several publications. Vermetids are well studied concerning the intraspecific competition with neighboring individuals but not in their interaction with other taxa like corals or fish. We did extensive in situ video recording and observed that fish avoided the plankton-load nets although several specialized taxa are known to be molluscivores, mucivores, and/or feed on plankton. As many molluscs use chemical weapons to combat feeding pressure and to defend themselves against predators, we screened empty and plankton-load mucus nets for potential bioactive metabolites. Bioactivity testing was performed with a recently developed system based on a chromatographic separation (high-performance thin-layer chromatography (HPTLC)) and a bioassay with luminescent bacteria Vibrio fischeri. Thus, we found at least two active compounds exclusively accumulated by the wormsnails themselves. This is the first record of bioactive properties in the whole family of Vermetidae

Unterrichtsmaterial für die Sekundarstufe I & II

Tittor A, Schwabe N, Center for Interamerican Studies (CIAS), eds. Menschen. Nutzen. Natur. Zum Umgang mit Rohstoffreichtum in Lateinamerika. Unterrichtsmaterialienreihe "Wissen um globale Verflechtungen". Vol 1. Bielefeld: kipu Verlag; 2016

Cognitive symptoms in a rat model of Parkinson's Disease

Objective: We here investigated the effect of bilateral 6-hydroxydopamine (6-OHDA) lesions, a rat model for Parkinson's disease (PD), on impulsivity and attention in an auditory oddball paradigm. In PD, the progressive loss of dopamine (DA) neurons in the substantia nigra leads to disturbed motor function, but cognitive disturbances, including attentional deficits and impulsivity, are increasingly recognized as disabling factors. Rats with 6-OHDA induced nigrostriatal lesions of dopamine neurons show significant motor impairment reminiscent of PD, and recent studies also indicate cognitive impairment in this model. Methods: Rats were trained in a 3-class auditory oddball paradigm, where they had to nose poke a hole after an infrequent correct tone, which was rewarded by a pellet, but to ignore a frequent standard tone and infrequent distractor tone. After reaching a criterion of 90% correct hits, retrograde degeneration of DA neurons in the substantia nigra were induced by bilateral striatal injection of 6-OHDA (10 μg in 1μl PBS; n=12), sham-lesioned rats (controls; n=8) received vehicle. Four weeks after surgery the rats were re-tested in the oddball paradigm. Results: After 6-OHDA lesions, rats show deteriorated attention, as indicated by a significant decrease in the hit rate to the correct tone. Additionally, the number of impulsive nose pokes was reduced compared to controls, which would indicate less impulsive behavior. Conclusion: We conclude that rats with bilateral 6-OHDA lesions may be used to investigate the biological basis of attentional deficits in PD, and to develop and test new therapeutic strategies for these symptoms ranging from pharmacological treatment to neurosurgical intervention

Reformulation of the D3(Becke–Johnson) Dispersion Correction without Resorting to Higher than <i>C</i>₆ Dispersion Coefficients

A reformulated version of Grimme’s most recent DFT dispersion correction with Becke–Johnson damping (DFT-D3­(BJ)) is presented, which only depends on <i>C</i>₆ dispersion coefficients. The role of the higher order correction terms in the DFT-D3­(BJ) model is critically investigated, and a sigmoidal interpolation function for adjusting to different density functional approximations (DFA) is employed alternatively, while keeping finite damping of Becke and Johnson. For the proposed <i>C</i>₆-only dispersion correction scheme, only one parameter needs to be fitted per DFA (instead of three for DFT-D3­(BJ)). Eight standard DFAs from different classes are parametrized and evaluated. In comparison to DFT-D3­(BJ), one of the most accurate corrections up to date, the new correction shows only negligible deviations in accuracy for the huge GMTKN30 benchmark set