C Hargrove

Acanthamoeba are free-living pathogenic protozoa that cause blinding keratitis, disseminated infection, and granulomatous amebic encephalitis, which is generally fatal. The development of efficient and safe drugs is a critical unmet need. Acanthamoeba sterol 14α-demethylase (CYP51) is an essential enzyme of the sterol biosynthetic pathway. Repurposing antifungal azoles for amoebic infections has been reported, but their inhibitory effects on Acanthamoeba CYP51 enzymatic activity have not been studied...

RNA-protein interactions are essential to RNA function throughout biology. Identifying the protein interactions associated with a specific RNA, however, is currently hindered by the need for RNA labeling or costly tiling-based approaches. Conventional strategies, which commonly rely on affinity pull-down approaches, are also skewed to the detection of high affinity interactions and frequently miss weaker interactions that may be biologically important. Reported here is the first adaptation of stability-based mass spectrometry methods for the global analysis of RNA-protein interactions...

The 14α-demethylation step is critical in eukaryotic sterol biosynthesis, catalyzed by cytochrome P450 (P450) Family 51 enzymes, e.g. with lanosterol in mammals. This conserved 3-step reaction terminates in a C-C cleavage step that generates formic acid, the nature of which has been controversial. Proposed mechanisms involve roles of P450 Compound 0 (ferric peroxide anion, FeO2¯) or Compound I (perferryl oxygen, FeO3+) reacting with either the aldehyde or its hydrate, respectively. Analysis of 18O incorporation into formic acid from 18O2 provides a means of distinguishing the two mechanisms...

S-adenosyl-L-methionine (SAM) is an abundant biomolecule used by methyltransferases to regulate a wide range of essential cellular processes such as gene expression, cell signaling, protein functions, and metabolism. Despite considerable effort, there remain many specificity challenges associated with designing small molecule inhibitors for methyltransferases, most of which exhibit off-target effects. Interestingly, NMR evidence suggests that SAM undergoes conformeric exchange between several states when free in solution...

Morphometric analysis of wings has been suggested for identifying and controlling isolated populations of tsetse (Glossina spp), vectors of human and animal trypanosomiasis in Africa. Single-wing images were captured from an extensive data set of field-collected tsetse wings of species Glossina pallidipes and G. m. morsitans. Morphometric analysis required locating 11 anatomical landmarks on each wing. The manual location of landmarks is time-consuming, prone to error, and infeasible for large data sets. We developed a two-tier method using deep learning architectures to classify images and make accurate landmark predictions...

Cytochrome P450 (P450, CYP) Family 51 enzymes catalyze the 14α-demethylation of sterols, leading to critical products used for membranes and the production of steroids, as well as signaling molecules. In mammals, P450 51 catalyzes the 3-step, 6-electron oxidation of lanosterol to form (4β,5α)-4,4-dimethyl-cholestra-8,14,24-trien-3-ol (FF-MAS). P450 51A1 can also use 24,25-dihydrolanosterol (a natural substrate in the Kandutsch-Russell cholesterol pathway). 24,25-Dihydrolanosterol and the corresponding P450 51A1 reaction intermediates, the 14α-alcohol and -aldehyde derivatives of dihydrolanosterol, were synthesized to study the kinetic processivity of the overall 14α-demethylation reaction of human P450 51A1...

The molecular evolution of cytochromes P450 and associated redox-driven oxidative catalysis remains a mystery in biology. It is widely believed that sterol 14α-demethylase (CYP51), an essential enzyme of sterol biosynthesis, is the ancestor of the whole P450 superfamily given its conservation across species in different biological kingdoms. Herein we have utilized X-ray crystallography, molecular dynamics simulations, phylogenetics and electron transfer measurements to interrogate the nature of P450-redox partner binding using the naturally occurring fusion protein, CYP51-ferredoxin found in the sterol-producing bacterium Methylococcus capsulatus...

The UBTF E210K neuroregression syndrome is caused by de novo dominant mutations in UBTF (NM_014233.3:c.628G > A, p.Glu210Lys). In humans, onset is typically at 2.5 to 3 years and characterized by slow progression of global motor, cognitive and behavioral dysfunction. Other potentially pathogenic UBTF variants have been reported in humans with severe neurological disease and it remains undetermined if the UBTF E210K mutation operates via gain- and/or loss-of-function. Here we examine the behavioral, cognitive, motor, and molecular effects of Ubtf knockout and knockdown in mice as a means of gauging the role of loss-of-function in humans...

Educational disparities in health are well documented, yet the education-health relationship is inconsistent across racial/ethnic and nativity groups. These inconsistencies may arise from characteristics of the early life environments in which individuals attain their education. We evaluate this possibility by investigating (1) whether educational disparities in cardiometabolic risk vary by race/ethnicity and nativity among Black, Hispanic, and White young adults; (2) the extent to which racial/ethnic-nativity differences in the education-health relationship are contingent on economic, policy, and social characteristics of counties of early life residence; and (3) the county characteristics associated with the best health at higher levels of education for each racial/ethnic-nativity group...

Most prosthetic limbs can autonomously move with dexterity, yet they are not perceived by the user as belonging to their own body. Robotic limbs can convey information about the environment with higher precision than biological limbs, but their actual performance is substantially limited by current technologies for the interfacing of the robotic devices with the body and for transferring motor and sensory information bidirectionally between the prosthesis and the user. In this Perspective, we argue that direct skeletal attachment of bionic devices via osseointegration, the amplification of neural signals by targeted muscle innervation, improved prosthesis control via implanted muscle sensors and advanced algorithms, and the provision of sensory feedback by means of electrodes implanted in peripheral nerves, should all be leveraged towards the creation of a new generation of high-performance bionic limbs...

Ozone exposure induces neuroendocrine stress response, which causes lymphopenia. It was hypothesized that ozone-induced increases in stress hormones will temporally follow changes in circulating granulocytes, monocytes and lymphocyte subpopulations. The goal of this study was to chronicle the changes in circulating stress hormones, cytokines, and leukocyte trafficking during 4 -h exposure to ozone. Male Wistar Kyoto rats were exposed to air or ozone (0.4 or 0.8 ppm) for 0.5, 1, 2, or 4 hours. After each time point, Circulating stress hormones and cytokines, and lung gene expression were assessed along with live and apoptotic granulocytes, monocytes (classical and non-classical), and lymphocytes (B, Th and Tc ) in blood, thymus and spleen using flow cytometry...

Methanobactins (MBs) are small (<1,300-Da) posttranslationally modified copper-binding peptides and represent the extracellular component of a copper acquisition system in some methanotrophs. Interestingly, MBs can bind a range of metal ions, with some being reduced after binding, e.g., Cu2+ reduced to Cu+ . Other metal ions, however, are bound but not reduced, e.g., K+ . The source of electrons for selective metal ion reduction has been speculated to be water but never empirically shown. Here, using H2 18 O, we show that when MBs from Methylocystis sp...

Unprecedented quantities of heat are entering the Pacific sector of the Arctic Ocean through Bering Strait, particularly during summer months. Though some heat is lost to the atmosphere during autumn cooling, a significant fraction of the incoming warm, salty water subducts (dives beneath) below a cooler fresher layer of near-surface water, subsequently extending hundreds of kilometers into the Beaufort Gyre. Upward turbulent mixing of these sub-surface pockets of heat is likely accelerating sea ice melt in the region...

The COVID-19 pandemic continues to strain health care systems and drive shortages in medical supplies and equipment around the world. Resource allocation in times of scarcity requires transparent, ethical frameworks to optimize decision making and reduce health care worker and patient distress. The complexity of allocating dialysis resources for both patients receiving acute and maintenance dialysis has not previously been addressed. Using a rapid, collaborative, and iterative process, BC Renal, a provincial network in Canada, engaged patients, doctors, ethicists, administrators, and nurses to develop a framework for addressing system capacity, communication challenges, and allocation decisions...

Bone regeneration is a critical area of research impacting treatment of diseases such as osteoporosis, age-related decline, and orthopaedic implants. A crucial question in bone regeneration is that of bone architectural quality, or how "good" is the regenerated bone tissue structurally? Current methods address typical long bone architecture, however there exists a need for improved ability to quantify structurally relevant parameters of bone in non-standard bone shapes. Here we present a new analysis approach based on open-source semi-automatic methods combining image processing, solid modeling, and numerical calculations to analyze bone tissue at a more granular level using μCT image data from a mouse digit model of bone regeneration...

Given the challenges to the notion of objectivity posed by social psychological research on investigator effects, constructivist and critical epistemological perspectives, and the introduction of qualitative research methods in psychology, the investigators examined how leading methodologists understand the function of objectivity and subjectivity in psychological science. The aim of the study was to learn how contemporary methodologists view these issues so as to communicate converging perspectives to the field and inform methods education...

Sterol biosynthesis, primarily associated with eukaryotic kingdoms of life, occurs as an abbreviated pathway in the bacterium Methylococcus capsulatus. Sterol 14α-demethylation is an essential step in this pathway and is catalyzed by cytochrome P450 51 (CYP51). In M. capsulatus, the enzyme consists of the P450 domain naturally fused to a ferredoxin domain at the C-terminus (CYP51fx). The structure of M. capsulatus CYP51fx was solved to 2.7 Å resolution and is the first structure of a bacterial sterol biosynthetic enzyme...

While across the animal kingdom offspring are born smaller than their parents, notable exceptions exist. Several dipteran species belonging to the Hippoboscoidea superfamily can produce offspring larger than themselves. In this essay, the blood-feeding tsetse is focused on. It is suggested that the extreme reproductive strategy of this fly is enabled by feeding solely on highly nutritious blood, and producing larval offspring that are soft and malleable. This immense reproductive expenditure may have evolved to avoid competition with other biting flies...

BACKGROUND AND AIMS: Following liver injury, mast cells (MCs) migrate into the liver and are activated in patients with cholestasis. Inhibition of MC mediators decreases ductular reaction (DR) and liver fibrosis. Transforming growth factor beta 1 (TGF-β1) contributes to fibrosis and promotes liver disease. Our aim was to demonstrate that reintroduction of MCs induces cholestatic injury through TGF-β1. APPROACH AND RESULTS: Wild-type, KitW-sh (MC-deficient), and multidrug resistance transporter 2/ABC transporter B family member 2 knockout mice lacking l-histidine decarboxylase were injected with vehicle or PKH26-tagged murine MCs pretreated with 0...

CYP51 enzymes (sterol 14α-demethylases) are cytochromes P450 that catalyze multistep reactions. The CYP51 reaction occurs in all biological kingdoms and is essential in sterol biosynthesis. It removes the 14α-methyl group from cyclized sterol precursors by first forming an alcohol, then an aldehyde, and finally eliminating formic acid with the introduction of a Δ14-15 double bond in the sterol core. The first two steps are typical hydroxylations, mediated by an electrophilic compound I mechanism. The third step, C-C bond cleavage, has been proposed to involve either compound I ( i...