Zachary Hudson

This study communicates physicians' barriers when counselling patients at different stages of PA behaviour change and the influence of career status on barrier experience. Developing patient-stage- and career-stage-specific medical training, interventions and policy changes may enhance PA counselling among physicians, and ultimately patient PA behaviour and health outcomes.Prolyl endopeptidases (PEPs) hydrolyze proteins to yield bioactive peptides and are effective in the treatment of celiac disease. However, the catalytic efficiency of PEPs still has the potential to be improved, which could further strengthen their industrial and therapeutic applications. Herein, a novel rational design strategy based on a "near-attack conformation" of the catalytic state of PEP was adopted. Constrained dynamic simulations were applied, followed by the virtual screening of potentially favorable mutants according to their binding free energy. We redesigned Sphaerobacter thermophiles PEP with high-temperature activity/stability, a wide range of pH stabilities, and high proline specificity. As a result, the kcat value of two PEP mutants (I462W and Q560Y) increased by 208.2 and 150.1%, respectively, and the kcat/KM increased by 32.7 and 6.3%, respectively. These data revealed that the PEP mutants had improved catalytic efficiency and that our strategy can be applied for enzyme engineering.Misregulation of the E3 ubiquitin ligase Parkin and the kinase PINK1 underlie both inherited and idiopathic Parkinson's disease-associated neurodegeneration. Parkin and PINK1 work together to catalyze the assembly of ubiquitin chains on substrates located on the outer mitochondrial membrane to facilitate autophagic removal of damaged mitochondria through a process termed mitophagy. Quantitative measurements of Parkin-mediated chain assembly, both in vitro and on mitochondria, have revealed that chains are composed of Lys6, Lys11, Lys48, and Lys63 linkages. The combinatorial nature of these chains is further expanded by the ability of PINK1 to phosphorylate individual subunits. The precise architecture of chains produced by the coordinated action of PINK1 and Parkin, however, are unknown. Here, we demonstrate that quantitative middle-down mass spectrometry using uniformly 15N-labeled ubiquitin variants as internal standards informs on the extent of chain branching. We find that Parkin is a prolific branching enzyme in vitro. Quantitative middle-down mass spectrometry also reveals that phospho-Ser65-ubiquitin (pSer65-Ub)-a key activator of Parkin-is not incorporated into chains to a significant extent. Our results suggest that Parkin-mediated chain branching is "on-pathway", and branch points are the principal targets of the deubiquitinase USP30.A detailed investigation was conducted on the reaction of a BINOL-coumarin-based fluorescent probe with amino acids. On the basis of the studies including fluorescence spectroscopy, 1H NMR, UV-Vis, mass spectroscopy, single crystal X-ray analysis and molecular modeling, it was found that the distinctively different fluorescent responses of the probe toward the amino acid at the two excitation wavelengths are due to two different reaction pathways that generate different intermediates and products.No data are available on the serum metabolomics and lipidomics profiles of people with asymptomatic intracranial arterial stenosis. We explored the characteristic metabolites of individuals with asymptomatic severe intracranial arterial stenosis (asICAS) using untargeted serum metabolomics and lipidomics analyses based on ultra-high-performance liquid chromatography high-resolution mass spectrometry (UPLC-HRMS). This case-control study included 25 participants with asICAS and 25 age- and sex-matched controls free of asICAS, who were all diagnosed by using magnetic resonance angiography and derived from the same population-based study. Serum metabolomics and lipidomics profiles were determined using UPLC-HRMS, and possible biomarker metabolites were identified. Compared with the control group, the asICAS group showed higher levels of free choline, glycerophosphocholine, uracil, taurine, and four peptide molecules and lower levels of free fatty acids, hydroxydodecanedioic acid, hydroxy valeryl carnitine, hydroxytetradecanedioic acid, and two sphingomyelin molecules. The serum metabolomics and lipidomics profiles for people with asICAS are characterized by abnormal metabolism of sphingomyelin, taurine/hypotaurine, pyrimidine, and protein (peptide). The biological changes in asICAS may mainly involve taurine/hypotaurine, glycerophospholipid, and sphingolipid metabolism pathways. Biofunctional analysis indicated that these differential metabolites were correlated with metabolic diseases such as early myocardial injury, heart failure, and diabetes.Phosphinidene complexes of a general formula RPM(CO)n (R = an alkyl or aryl group; M = a transition metal) are electrophilic and thermally unstable. Thus, the isolation of these elusive species for structural elucidations remains so far a challenge. Herein, we report the first terminal phosphinidene complexes [(NHC)C(Ph)P]Fe(CO)4 (NHC = IPr = C(NDipp)CH2, 3; Me-IPr = C(NDipp)CMe2, 4; Dipp = 2,6-iPr2C6H3; NHC = N-heterocyclic carbene) as red crystalline solids containing a π-donor N-heterocyclic vinyl (NHV) substituent at the phosphorus atom. Calculations reveal the donor-acceptor type bonding between phosphorus and iron atoms in 3 and 4. The P→Fe donation represents ~70 % whereas the Fe→P π-back donation corresponds to ~15 % of the orbital interaction. The phosphorus atom in 3 and 4 carries +0.65e and +0.64e charge, respectively, indicating an electrophilic character of the phosphinidene (NHC)C(Ph)P moiety. Selleckchem DL-Thiorphan Accordingly, 3 reacts with an NHC nucleophile (IMe4) to yield the Lewis adduct [(NHC)C(Ph)P(IMe4)]Fe(CO)4 (5) (IMe4 = C(NMeCMe)2). The coordination of an electron-rich NHC (IMe4) to the phosphorus atom in 5 precludes the π-electron density transfer from the NHV to the phosphorus atom. Thus, the CIPr‒CVinyl and CVinyl‒P bonds of 5 become respectively shorter and longer compared to those of 3.