Two consecutive cruises in the Weddell Sea, Antarctica, in winter 2013 provided the first direct observations of sea salt aerosol (SSA) production from blowing snow above sea ice, thereby validating ...a model hypothesis to account for winter time SSA maxima in the Antarctic. Blowing or drifting snow often leads to increases in SSA during and after storms. For the first time it is shown that snow on sea ice is depleted in sulfate relative to sodium with respect to seawater. Similar depletion in bulk aerosol sized ∼0.3–6 µm above sea ice provided the evidence that most sea salt originated from snow on sea ice and not the open ocean or leads, e.g. >90 % during the 8 June to 12 August 2013 period. A temporally very close association of snow and aerosol particle dynamics together with the long distance to the nearest open ocean further supports SSA originating from a local source. A mass budget estimate shows that snow on sea ice contains even at low salinity (<0.1 psu) more than enough sea salt to account for observed increases in atmospheric SSA during storms if released by sublimation. Furthermore, snow on sea ice and blowing snow showed no or small depletion of bromide relative to sodium with respect to seawater, whereas aerosol was enriched at 2 m and depleted at 29 m, suggesting that significant bromine loss takes place in the aerosol phase further aloft and that SSA from blowing snow is a source of atmospheric reactive bromine, an important ozone sink, even during winter darkness. The relative increase in aerosol concentrations with wind speed was much larger above sea ice than above the open ocean, highlighting the importance of a sea ice source in winter and early spring for the aerosol burden above sea ice. Comparison of absolute increases in aerosol concentrations during storms suggests that to a first order corresponding aerosol fluxes above sea ice can rival those above the open ocean depending on particle size. Evaluation of the current model for SSA production from blowing snow showed that the parameterizations used can generally be applied to snow on sea ice. Snow salinity, a sensitive model parameter, depends to a first order on snowpack depth and therefore was higher above first-year sea ice (FYI) than above multi-year sea ice (MYI). Shifts in the ratio of FYI and MYI over time are therefore expected to change the seasonal SSA source flux and contribute to the variability of SSA in ice cores, which represents both an opportunity and a challenge for the quantitative interpretation of sea salt in ice cores as a proxy for sea ice.
Post-translational modification catalyzed by phosphopantetheinyl transferases (PPTases) has previously been used to site-specifically label proteins with structurally diverse molecules. PPTase ...catalysis results in covalent modification of a serine residue in acyl/peptidyl carrier proteins and their surrogate substrates which are typically fused to the N- or C-terminus. To test the utility of PPTases for preparing antibody–drug conjugates (ADCs), we inserted 11 and 12-mer PPTase substrate sequences at 110 constant region loop positions of trastuzumab. Using Sfp-PPTase, 63 sites could be efficiently labeled with an auristatin toxin, resulting in 95 homogeneous ADCs. ADCs labeled in the CH1 domain displayed in general excellent pharmacokinetic profiles and negligible drug loss. A subset of CH2 domain conjugates underwent rapid clearance in mouse pharmacokinetic studies. Rapid clearance correlated with lower thermal stability of the particular antibodies. Independent of conjugation site, almost all ADCs exhibited subnanomolar in vitro cytotoxicity against HER2-positive cell lines. One selected ADC was shown to induce tumor regression in a xenograft model at a single dose of 3 mg/kg, demonstrating that PPTase-mediated conjugation is suitable for the production of highly efficacious and homogeneous ADCs.
In vivo incorporation of isotopically labeled unnatural amino acids into large proteins drastically reduces the complexity of nuclear magnetic resonance (NMR) spectra. Incorporation is accomplished ...by coexpressing an orthogonal tRNA/aminoacyl−tRNA synthetase pair specific for the unnatural amino acid added to the media and the protein of interest with a TAG amber codon at the desired incorporation site. To demonstrate the utility of this approach for NMR studies, 2-amino-3-(4-(trifluoromethoxy)phenyl)propanoic acid (OCF3Phe), 13C/15N-labeled p-methoxyphenylalanine (OMePhe), and 15N-labeled o-nitrobenzyl-tyrosine (oNBTyr) were incorporated individually into 11 positions around the active site of the 33 kDa thioesterase domain of human fatty acid synthase (FAS-TE). In the process, a novel tRNA synthetase was evolved for OCF3Phe. Incorporation efficiencies and FAS-TE yields were improved by including an inducible copy of the respective aminoacyl-tRNA synthetase gene on each incorporation plasmid. Using only between 8 and 25 mg of unnatural amino acid, typically 2 mg of FAS-TE, sufficient for one 0.1 mM NMR sample, were produced from 50 mL of Escherichia coli culture grown in rich media. Singly labeled protein samples were then used to study the binding of a tool compound. Chemical shift changes in 1H−15N HSQC, 1H−13C HSQC, and 19F NMR spectra of the different single site mutants consistently identified the binding site and the effect of ligand binding on conformational exchange of some of the residues. OMePhe or OCF3Phe mutants of an active site tyrosine inhibited binding; incorporating 15N-Tyr at this site through UV-cleavage of the nitrobenzyl-photocage from oNBTyr re-established binding. These data suggest not only robust methods for using unnatural amino acids to study large proteins by NMR but also establish a new avenue for the site-specific labeling of proteins at individual residues without altering the protein sequence, a feat that can currently not be accomplished with any other method.
Uncertainty and interconnectedness in complex engineering and engineered systems such as power-grids and telecommunication networks are sources of vulnerability compromising the resilience of these ...systems. Conditions of uncertainty and interconnectedness change over time and depend on emerging socio-technical contexts, thus conventional methods which can conduct normative, descriptive and prescriptive assessment of complex engineering and engineered systems resilience are limited. This paper brings together contributions of experts in complex engineering and engineered systems who have identified six methods, three each for uncertainty and interconnectedness, which form the foundational methods for knowing complex engineering and engineered systems resilience. The paper has reviewed how these methods contribute to overcoming uncertainty or interconnectedness and how they are implemented using case studies in order to illustrate essential approaches to enhancing resilience. It is hoped that this approach will allow the subject to be quantified and best practice standards to develop.
The goal of this study was to determine the intra- and interday reliability of pressure pain thresholds (PPT) in the upper extremity and torso of asymptomatic women. Nineteen healthy women (20-39 ...years) with no underlying musculoskeletal problems had 3 PPT trials performed on 8 different locations in the upper extremity and torso over 4 consecutive days. The test-retest reliability of PPT values was robust and highly consistent over the 4 days. The PPT intraclass correlations (ICC) were highly consistent and repeatable over the 4 days of testing (day 1: ICC = 0.94; day 2: ICC = 0.96; day 3: ICC = 0.97 and day 4: ICC = 0.96). When compared with baseline measurements obtained on day 1, the PPT values were significantly lower (P < .05) on days 2, 3, and 4 at all 8 locations. Although the PPT test-retest reliability is robust and consistent throughout the 4 days, there appears to be a similar overall decline in the magnitude of the absolute PPT response at each of the 8 locations. A specific explanation for this greater overall sensitivity in PPTs at all 8 locations is lacking; however, a centrally mediated alteration in pressure/pain sensation could contribute to the overall trend observed in this study.
PPT measurements of the upper limb and torso will be significantly lower with repeated measures over a short period time. A standardized evaluation grid should be included in baseline so as to accurately evaluate the progression in shoulder rehabilitation in women with shoulder dysfunction.
Tuberculosis continues to be a global health threat, making bicyclic nitroimidazoles an important new class of therapeutics. A deazaflavin-dependent nitroreductase (Ddn) from Mycobacterium ...tuberculosis catalyzes the reduction of nitroimidazoles such as PA-824, resulting in intracellular release of lethal reactive nitrogen species. The N-terminal 30 residues of Ddn are functionally important but are flexible or access multiple conformations, preventing structural characterization of the full-length, enzymatically active enzyme. Several structures were determined of a truncated, inactive Ddn protein core with and without bound F420 deazaflavin coenzyme as well as of a catalytically competent homolog from Nocardia farcinica. Mutagenesis studies based on these structures identified residues important for binding of F420 and PA-824. The proposed orientation of the tail of PA-824 toward the N terminus of Ddn is consistent with current structure-activity relationship data.
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► Crystal structures of Ddn, the reductase for antitubercular drug PA-824 ► Active site residues and F420-binding mode described ► The N terminus is important for PA-824 binding but could not be fully characterized ► An active homolog from Nocardia farcinica was identified and studied
Symptoms may persist after the initial phases of COVID-19 infection, a phenomenon termed long COVID. Current knowledge on long COVID has been mostly derived from test-confirmed and hospitalized ...COVID-19 patients. Data are required on the burden and predictors of long COVID in a broader patient group, which includes both tested and untested COVID-19 patients in primary care.
This is an observational study using data from Platform C19, a quality improvement program-derived research database linking primary care electronic health record data (EHR) with patient-reported questionnaire information. Participating general practices invited consenting patients aged 18-85 to complete an online questionnaire since 7th August 2020. COVID-19 self-diagnosis, clinician-diagnosis, testing, and the presence and duration of symptoms were assessed via the questionnaire. Patients were considered present with long COVID if they reported symptoms lasting ≥4 weeks. EHR and questionnaire data up till 22nd January 2021 were extracted for analysis. Multivariable regression analyses were conducted comparing demographics, clinical characteristics, and presence of symptoms between patients with long COVID and patients with shorter symptom duration.
Long COVID was present in 310/3151 (9.8%) patients with self-diagnosed, clinician-diagnosed, or test-confirmed COVID-19. Only 106/310 (34.2%) long COVID patients had test-confirmed COVID-19. Risk predictors of long COVID were age ≥40 years (adjusted Odds Ratio AdjOR=1.49 1.05-2.17), female sex (adjOR=1.37 1.02-1.85), frailty (adjOR=2.39 1.29-4.27), visit to A&E (adjOR=4.28 2.31-7.78), and hospital admission for COVID-19 symptoms (adjOR=3.22 1.77-5.79). Aches and pain (adjOR=1.70 1.21-2.39), appetite loss (adjOR=3.15 1.78-5.92), confusion and disorientation (adjOR=2.17 1.57-2.99), diarrhea (adjOR=1.4 1.03-1.89), and persistent dry cough (adjOR=2.77 1.94-3.98) were symptom features statistically more common in long COVID.
This study reports the factors and symptom features predicting long COVID in a broad primary care population, including both test-confirmed and the previously missed group of COVID-19 patients.
Objectives
Children who have been maltreated are at increased risk of further maltreatment. Identification of those at highest risk of further maltreatment is a priority for professionals working in ...child protection services. The current study is intended to consolidate and expand on previous work on recurrence of child maltreatment. It has sought to identify risk factors for maltreatment recurrence in the recent literature in the expectation that this may help in the practical identification of children at risk.
Methods
We conducted a systematic review of cohort studies published between 2003 and 2009, identifying factors associated with maltreatment recurrence in children. Studies included demonstrated differing levels of substantiation of maltreatment.
Results
Fifteen studies met inclusion criteria but showed significant heterogeneity, varying in setting, recruitment of subjects, types of maltreatment considered and length of follow-up. Previous findings were replicated and expanded in the current study in relation to a range of factors, including rates of maltreatment recurrence, maltreatment types, frequency of previous episodes of maltreatment, child and family considerations, home environment and service provision. Factors were identified irrespective of level of maltreatment substantiation.
Conclusion
This study provides further systematic evidence of the existence of a number of factors associated with child maltreatment recurrence. It points to the possibility of practical application of its findings within the wider context of decision making in child protection services, with the ultimate aim of reducing recurrence of maltreatment in individual cases.
Pyrroline-carboxy-lysine (Pcl) is a demethylated form of pyrrolysine that is generated by the pyrrolysine biosynthetic enzymes when the growth media is supplemented with D-ornithine. Pcl is readily ...incorporated by the unmodified pyrrolysyl-tRNA/tRNA synthetase pair into proteins expressed in Escherichia coli and in mammalian cells. Here, we describe a broadly applicable conjugation chemistry that is specific for Pcl and orthogonal to all other reactive groups on proteins. The reaction of Pcl with 2-amino-benzaldehyde or 2-amino-acetophenone reagents proceeds to near completion at neutral pH with high efficiency. We illustrate the versatility of the chemistry by conjugating Pcl proteins with poly(ethylene glycol)s, peptides, oligosaccharides, oligonucleotides, fluorescence, and biotin labels and other small molecules. Because Pcl is genetically encoded by TAG codons, this conjugation chemistry enables enhancements of the pharmacology and functionality of proteins through site-specific conjugation.
Much research has shown that spouses of combat veterans with posttraumatic stress disorder (PTSD) have higher rates of psychological and marital distress than do spouses of veterans without PTSD; ...however, very few studies have examined potential mechanisms of this increased vulnerability. The current study examined spouses of National Guard soldiers recently returned from deployments in Iraq. In addition to documenting elevated levels of psychological symptoms in these spouses, the authors found that spouses experienced greater symptom severity when they perceived high levels of symptoms in soldiers but the soldiers endorsed low levels of symptoms. Furthermore, spouses' marital satisfaction was negatively linked to soldiers' self-reported symptom severity only when spouses perceived that soldiers had experienced low levels of combat activity while deployed. When spouses perceived high levels of such activity, soldiers' self-reported symptoms had no relationship with spouses' marital satisfaction. These findings highlight the importance of interpersonal perceptions in intimate relationships and are consistent with the notion that uncontrollable attributions for a relative's mental health problems may provide a buffer against relationship distress.
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