We have initiated a broad-based program aimed at understanding the molecular basis of fluorine specificity in enzymatic systems, and in this context, we report crystallographic and biochemical ...studies on a fluoroacetyl-coenzyme A (CoA) specific thioesterase (FlK) from Streptomyces cattleya. Our data establish that FlK is competent to protect its host from fluoroacetate toxicity in vivo and demonstrate a 106-fold discrimination between fluoroacetyl-CoA (k cat/K M = 5 × 107 M−1 s−1) and acetyl-CoA (k cat/K M = 30 M−1 s−1) based on a single fluorine substitution that originates from differences in both substrate reactivity and binding. We show that Thr 42, Glu 50, and His 76 are key catalytic residues and identify several factors that influence substrate selectivity. We propose that FlK minimizes interaction with the thioester carbonyl, leading to selection against acetyl-CoA binding that can be recovered in part by new CO interactions in the T42S and T42C mutants. We hypothesize that the loss of these interactions is compensated by the entropic driving force for fluorinated substrate binding in a hydrophobic binding pocket created by a lid structure, containing Val 23, Leu 26, Phe 33, and Phe 36, that is not found in other structurally characterized members of this superfamily. We further suggest that water plays a critical role in fluorine specificity based on biochemical and structural studies focused on the unique Phe 36 “gate” residue, which functions to exclude water from the active site. Taken together, the findings from these studies offer molecular insights into organofluorine recognition and design of fluorine-specific enzymes.
Autism is characterized by restricted, repetitive behaviors and impairment in socialization and communication. Although no neuropathologic substrate underlying autism has been found, the findings of ...brain overgrowth via neuroimaging studies and increased levels of brain-derived neurotrophic factor (BDNF) in neuropathologic and blood studies favor an anabolic state. We examined acetylcholinesterase, plasma neuronal proteins, secreted beta-amyloid precursor protein (APP), and amyloid-beta 40 and amyloid-beta 42 peptides in children with and without autism. Children with severe autism and aggression expressed secreted beta-amyloid precursor protein at two or more times the levels of children without autism and up to four times more than children with mild autism. There was a trend for children with autism to show higher levels of secreted beta-amyloid precursor protein and nonamyloidogenic secreted beta-amyloid precursor protein and lower levels of amyloid-beta 40 compared with controls. This favors an increased α-secretase pathway in autism (anabolic), opposite to what is seen in Alzheimer disease. Additionally, a complex relationship between age, acetylcholinesterase, and plasma neuronal markers was found. (J Child Neurol 2006;21:444—449; DOI 10.2310/7010.2006.00130).
This study represents the first airborne, in-situ measurements of HC(O)OH in the European Arctic, across the winter and summer seasons. HC(O)OH concentrations are under predicted at present, ...particularly in the mid to high northern latitudes. Data presented here probe unconfirmed sources of HC(O)OH in the Arctic, and would suggest an ocean source of HC(O)OH is more significant than proposed land sources in both winter and summer environments. A maximum concentration of 420 ppt was recorded over the ocean during the July 2012 campaign. This was more than 1.7 times greater than the maximum land concentration reported. Calculated estimates on HC(O)OH production would suggest diiodomethane photolysis could represent a significant source of HC(O)OH in marine environments in the European Arctic. Enhanced HC(O)OH concentrations observed at altitudes greater than 2 km particularly during the March campaign highlight the significance of long range transport on the European Arctic budget. In addition, two HC(O)OH vertical profiles between the altitudes 0.3–6.6 km are presented to provide a more representative vertical profile for this latitude which may be used to improve forthcoming regional and global modelling of the HC(O)OH budget.
•The first airborne measurements of HC(O)OH in the European Arctic (EA).•No significant HC(O)OH snowpack source was observed during winter measurements.•Averaged HC(O)OH concentrations suggest marine sources dominate over land sources.•Calculations suggest CH2I2 oxidation represents a major marine source in the EA.
Tumor protein D52 (also known as CRHSP-28) is highly expressed in multiple cancers and tumor-derived cell lines; however, it is normally abundant in secretory epithelia throughout the digestive ...system, where it has been implicated in Ca(2+)-dependent digestive enzyme secretion (41). Here we demonstrate, using site-specific mutations, that Ca(2+)-sensitive phosphorylation at serine 136 modulates the accumulation of D52 at the plasma membrane within 2 min of cell stimulation. When expressed in Chinese hamster ovary CHO-K1 cells, D52 colocalized with adaptor protein AP-3, Rab27A, vesicle-associated membrane protein VAMP7, and lysosomal-associated membrane protein LAMP1, all of which are present in lysosome-like secretory organelles. Overexpression of D52 resulted in a marked accumulation of LAMP1 on the plasma membrane that was further enhanced following elevation of cellular Ca(2+). Strikingly, mutation of serine 136 to alanine abolished the Ca(2+)-stimulated accumulation of LAMP1 at the plasma membrane whereas phosphomimetic mutants constitutively induced LAMP1 plasma membrane accumulation independent of elevated Ca(2+). Identical results were obtained for endogenous D52 in normal rat kidney and HeLA cells, where both LAMP1 and D52 rapidly accumulated on the plasma membrane in response to elevated cellular Ca(2+). Finally, D52 induced the uptake of LAMP1 antibodies from the cell surface in accordance with both the level of D52 expression and phosphorylation at serine 136 demonstrating that D52 altered the plasma membrane recycling of LAMP1-associated secretory vesicles. These findings implicate both D52 expression and Ca(2+)-dependent phosphorylation at serine 136 in lysosomal membrane trafficking to and from the plasma membrane providing a novel Ca(2+)-sensitive pathway modulating the lysosome-like secretory pathway.
Mark Stockman: Evangelist for Plasmonics Aizpurua, Javier; Atwater, Harry A; Baumberg, Jeremy J ...
ACS photonics,
03/2021, Letnik:
8, Številka:
3
Journal Article
Odprti dostop
Mark Stockman was a founding member and evangelist for the plasmonics field for most of his creative life. He never sought recognition, but fame came to him in a different way. He will be dearly ...remembered by colleagues and friends as one of the most influential and creative contributors to the science of light from our generation.
The clustered regularly interspaced short palindromic repeats (CRISPR)-associated enzyme Cas9 is an RNA-guided endonuclease that uses RNA-DNA base-pairing to target foreign DNA in bacteria. ...Cas9-guide RNA complexes are also effective genome engineering agents in animals and plants. Here we use single-molecule and bulk biochemical experiments to determine how Cas9-RNA interrogates DNA to find specific cleavage sites. We show that both binding and cleavage of DNA by Cas9-RNA require recognition of a short trinucleotide protospacer adjacent motif (PAM). Non-target DNA binding affinity scales with PAM density, and sequences fully complementary to the guide RNA but lacking a nearby PAM are ignored by Cas9-RNA. Competition assays provide evidence that DNA strand separation and RNA-DNA heteroduplex formation initiate at the PAM and proceed directionally towards the distal end of the target sequence. Furthermore, PAM interactions trigger Cas9 catalytic activity. These results reveal how Cas9 uses PAM recognition to quickly identify potential target sites while scanning large DNA molecules, and to regulate scission of double-stranded DNA. PUBLICATION ABSTRACT
Background
Transient hypothyroxinaemia of prematurity (THOP) is associated with increased risk of cerebral palsy and lower IQ in low‐birthweight infants. This study explores whether THOP is also ...associated with increased risk of autism spectrum disorders (ASD).
Methods
This secondary analysis uses data from a birth cohort of newborns weighing 500 –2000 g (n = 1105) who were followed to age 21 years, when they were assessed for ASD in the second of a two‐stage process. Of the 187 assessed at age 21, 14 had ASD. Neonatal thyroxine results were available for 12/14 and 165/173 participants diagnosed with and without ASD, respectively. THOP was defined as thyroxine z‐score <−2.6. Unadjusted relative risks (RR) and confidence intervals (CI) were calculated.
Results
The mean neonatal thyroxine z‐score in young adults diagnosed with ASD was 0.5 SD lower 95% CI −0.16, 1.06 than in those without ASD. Participants with THOP were at 2.5‐fold greater risk of ASD (RR 2.5 95% CI 0.7, 8.4). While neither of these differences was statistically significant, in a secondary subgroup analysis of those whose mothers did not have hypertension during pregnancy, THOP significantly increased the RR for ASD (5.0 95% CI 1.2, 20.5).
Conclusion
While the primary relation between THOP and ASD found here is not statistically significant, the magnitude of association and significant relationship observed in the subgroup whose mothers did not have hypertension during pregnancy suggest that it is worthy of further investigation.
Drilling predation is among the most studied biotic interactions in the fossil record, and its overall patterns are well established on Cenozoic mollusks from North America. Few studies have examined ...such predation in Europe, which experienced a different geologic history. This study aims to evaluate taxonomic and environmental effects on molluscan drilling intensities from the Miocene of the Central Paratethys using drill frequency (DF) and prey effectiveness (PE), a measure of prey's ability to survive predatory attacks. 166 bulk samples from Austria and Slovakia that included 39,234 whole shells from the Karpatian (upper Burdigalian) and Badenian (Langhian and lower Serravallian) showed that at the level of stages, environments and localities, DF and PE were always below 10% and were slightly higher in bivalves than gastropods. Predation intensities from the Central Paratethys are therefore distinctly lower than those of other Miocene seas and it is hypothesized that this is explained by the rarity of naticid and muricid gastropods in the study area. The underlying factors controlling abundances of these drilling predators in the Central Paratethys could be related to the complex paleogeographic history of this inland sea. Intertidal DFs and PEs increased temporally, but differences in sublittoral DFs were not significant and PE values decreased significantly. Temporal patterns in DF and PE are influenced by disparities in sampled environments between the lower and middle Miocene. In general, comparisons of DFs across environments within the Karpatian and Badenian yielded similar results using higher and lower taxa. In contrast, such comparisons of PE were dependent upon taxonomic resolution.
The etiology of dementia that occurs in patients with schizophrenia is not well understood. Nicotinic acetylcholine receptors have been implicated in cognitive function, and deficits in these ...receptors have been reported in schizophrenia.
The present study investigates possible associations of nicotinic receptor subunit expression in the dorsal lateral prefrontal cortex, an area known to be affected in schizophrenia, and dementia rating.
α7 immunoreactivity was reduced by 20% to 28% and
3Hepibatidine binding was increased twofold in groups of patients with schizophrenia compared to normal control subjects matched for age, postmortem delay, and low levels of brain nicotine and cotinine. In contrast, no significant differences in α4, α3, or β2 immunoreactivity or α7 messenger RNA expression were observed in schizophrenia patients compared with control subject values. Clinical dementia ratings in patients with schizophrenia were correlated with neither
3Hepibatidine binding nor nicotinic receptor subunit expression.
These data indicate no relationship between the trend for reduced neocortical α7 subunit protein expression in schizophrenia and dementia. Further investigations are required to establish whether the reduction in α7 protein in the dorsal lateral prefrontal cortex is associated with clinical features other than dementia in schizophrenia.
Objectives: Affect and loneliness are important indicators of mental health and well-being in older adulthood and are linked to significant outcomes including physical health and mortality. Given a ...large focus on young-old adults within gerontological research, the primary aim of this study was to examine the ability of individual and social resources in predicting affect and loneliness within a sample of oldest-old individuals including centenarians, an understudied population.
Methods: Participants were assessed during the most recent cross-sectional data collection of the Georgia Centenarian Study. The eligible sample included 55 octogenarians (M = 83.70 years, SD = 2.68; range = 81-90) and 77 centenarians (M = 99.78 years, SD = 1.64; range = 98-109). Subjects scored 17 or greater on the Mini-Mental Status Exam and completed mental health assessments.
Results: Hierarchical regression analyses were conducted to examine the relation of affect and loneliness with demographic characteristics, physical and social functioning, cognition, and personality. Within this sample of cognitively intact oldest old, measures of executive control and cognitive functioning demonstrated limited association with mental health. Personality, specifically neuroticism, was strongly related to mental health indicators for both age groups and social relations were particularly important associates of centenarians' mental health.
Discussion: Findings indicate the distinctiveness of mental health indicators and the need to distinguish differential roles of individual and social resources in determining these outcomes among octogenarians and centenarians
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