Chemical genetic inhibition of DEAD-box proteins using covalent complementarity.

DEAD-box proteins are an essential class of enzymes involved in all stages of RNA metabolism. The study of DEAD-box proteins is challenging in a native setting since they are structurally similar, often essential and display dosage sensitivity. Pharmacological inhibition would be an ideal tool to probe the function of these enzymes. In this work, we describe a chemical genetic strategy for the specific inactivation of individual DEAD-box proteins with small molecule inhibitors using covalent complementarity. We identify a residue of low conservation within the P-loop of the nucleotide-binding site of DEAD-box proteins and show that it can be mutated to cysteine without a substantial loss of enzyme function to generate electrophile-sensitive mutants. We then present a series of small molecules that rapidly and specifically bind and inhibit electrophile-sensitive DEAD-box proteins with high selectivity over the wild-type enzyme. Thus, this approach can be used to systematically generate small molecule-sensitive alleles of DEAD-box proteins, allowing for pharmacological inhibition and functional characterization of members of this enzyme family.

Glycosylation of a model proto-RNA nucleobase with non-ribose sugars: implications for the prebiotic synthesis of nucleosides.

The emergence of nucleosides is an important, but poorly understood, element of the origins of life. We show that 2,4,6-triaminopyrimidine (TAP), a possible ancestral nucleobase of RNA, is glycosylated in water by non-ribose sugars in yields comparable to those previously reported for its reaction with ribose. The various sugars surveyed include ketoses and aldoses; tetroses, pentoses, and hexoses and are neutral, anionic, or cationic. Though they vary greatly in structure and properties, the data show that all sugars tested form glycosides with TAP. The structures of the eight TAP glycosides formed with glucose and two of its derivatives, glucose-6-phosphate and N-acetylglucosamine, were found to be ß-pyranosides with the glycosylation site on TAP varying with sugar identity. Our results suggest that prebiotic nucleoside formation would not have been restricted to ribose if ancestral RNA (or proto-RNA) utilized TAP and/or other proto-nucleobases with similar reactivities, and that the ability to form higher-order structures may have influenced proto-RNA monomer selection.

Adapting forensic case reporting to account for marginalization and vulnerability.

Scholarship of forensic sciences has shown politicalization of human remains and potential biases in criminal investigations. Specifically, concerns have been raised regarding how forensic anthropology analysis and documentation may hinder identification processes or obfuscate other data. As part of this scholarship, some have suggested that forensic anthropologists expand their reporting to include broader public health and safety information as well as reconsider who should be included in reports of anthropological findings. In response to these burgeoning discussions, this piece provides examples of ways anthropologists may formulate reports that capture evidence of marginalization or structural vulnerability. Documentation of findings can occur in myriad formats, including, but not limited to, individual case reports, reports on population analyses from cases, collaborative end-of-year reporting conducted with other medicolegal professionals, and collaborative databasing. This piece provides various templates and suggestions for reporting this kind of data while encouraging further discussion on related merits and concerns.

Defining the Culture of Surgery.

BACKGROUND: Surgical culture is often referenced but not well defined. Recent research and changing policies in graduate medical education have influenced the training paradigm and expectations of surgical trainees. It is unclear how these changes impact surgeons' understanding of surgical culture today and how those views impact surgical training. We sought to understand surgical culture and its impact on training from the perspective of a diverse group of surgeons with varied amounts of experience. STUDY DESIGN: A series of semi-structured, qualitative interviews were conducted with 21 surgeons and trainees in a single academic institution. Interviews were transcribed, coded, and analyzed using directed content analysis. RESULTS: We identified 7 major themes that impact surgical culture. Cohorts were separated by those who had been promoted to at least associate professor (late-career surgeons) and assistant professors, fellows, residents, and students (early-career surgeons). Both cohorts similarly emphasized patient-centered care, hierarchy, high standards, and meaningful work. Late- and early-career surgeons highlighted themes differently: late-career surgeons' perceptions were informed by experience and focused on challenges, complications, humility, and work ethic, while perceptions of early-career surgeons were more individually focused and referenced being goal-oriented and self-sacrificing, and focus on education and work-life balance. CONCLUSIONS: Late- and early-career surgeons both emphasize that patient-centered care is core to surgical culture. Early-career surgeons expressed more themes related to personal well-being, while late-career surgeons emphasized themes related to professional accomplishment. Differences in the perceived culture can lead to strained interactions between generations of surgeons and trainees, and a better understanding of these differences would lead to improved communication and interactions between these groups, as well as better management of expectations for surgeons in their training and career.

Recommendations for large-scale exhumations of Potter's Fields: Cases in the US.

Increased commitment to humanitarian identification and advances in DNA technology contribute to growing interest in large scale identification operations for unidentified, interred remains. However, scant literature discusses the practical challenges in conducting cemetery exhumations en masse for the purposes of DNA sampling and reinterment. This paper draws on a case study of cemetery exhumations conducted in the US as part of a multi-agency, multi-year endeavor, Operation UNITED. Challenges and strategies for managing logistics and manpower, locating human remains in active cemeteries, and overcoming containment and preservation issues of adult and juvenile remains are presented. This paper evidences the potential for investigation into cemetery exhumation strategies to assist in effective humanitarian identification.

IFITM proteins assist cellular uptake of diverse linked chemotypes.

The search for cell-permeable drugs has conventionally focused on low-molecular weight (MW), nonpolar, rigid chemical structures. However, emerging therapeutic strategies break traditional drug design rules by employing flexibly linked chemical entities composed of more than one ligand. Using complementary genome-scale chemical-genetic approaches we identified an endogenous chemical uptake pathway involving interferon-induced transmembrane proteins (IFITMs) that modulates the cell permeability of a prototypical biopic inhibitor of MTOR (RapaLink-1, MW: 1784 g/mol). We devised additional linked inhibitors targeting BCR-ABL1 (DasatiLink-1, MW: 1518 g/mol) and EIF4A1 (BisRoc-1, MW: 1466 g/mol), uptake of which was facilitated by IFITMs. We also found that IFITMs moderately assisted some proteolysis-targeting chimeras and examined the physicochemical requirements for involvement of this uptake pathway.

Astertarone A Synthase from Chinese Cabbage Does Not Produce the C4-Epimer: Mechanistic Insights.

The Brassica rapa oxidosqualene cyclase Bra032185 makes 60% astertarone A (1) and 20 minor triterpenes (0.1-11%). Mechanistic analysis indicates the absence of an enol intermediate to generate the 4 S epimer of 1, unless workup involves saponification. Bra032185 and its closest Arabidopsis thaliana homologue diverged markedly in product profiles over a short evolutionary distance, while developing opposite C18 configurations. Active-site residue comparison of Bra032185 with friedelin and shionone synthases suggests convergent evolution to 3-ketotriterpenes.

Metric sex estimation from the postcranial skeleton for the Colombian population.

This research explores the best univariate and multivariate indicators for sex estimation using 51 standard osteometric measurements of all six major postcranial long bones, bones of the shoulder girdle, pelvic girdle, and the calcaneus from a modern, Colombian skeletal collection. The hypotheses being tested are (1) that postcrania will yield accurate sex classification rates and (2) the shoulder girdle will demonstrate the highest discrimination, based on results from previous research. The sample consists of 134 individuals (50 females, 84 males) between the ages of 19 and 93 with a mean age of 47 years. The sample is from the Colombian Skeletal Collection, consisting of skeletons from cemeteries in Bogotá, with recent years of death. The methods include univariate and multivariate discriminant function analysis (DFA). The results for this sample indicate the same general pattern of univariate classification effectiveness as found in research on North Americans; however, here the humerus performs better than the distal femur and proximal tibia as demonstrated in some North American samples. The cross-validated percent correct univariate classification for the postcranial elements ranges from 64.8% to 86.1% (p<0.05). The highest univariate classification rates were for the scapular height (86.1%) and the humeral head diameter (86.0%). The highest multivariate classification rates are with the scapula (93.5%), os coxa (92.0%), clavicle (89.9%), humerus (89.1%), and ulna (89.1%). This study provides sectioning points to easily estimate sex using Colombian population-specific formulae. This research aids in forensic individuation, as the long bones of the postcranial elements are relatively resistant to taphonomic processes. Furthermore, the ability to achieve such a high degree of success from a single bone is preferable for the fast-paced forensic anthropology laboratories in Colombia that process hundreds to thousands of cases each year. This research plays an important role in the development of population standards in Colombia and South America and provides a robust method that can withstand courtroom scrutiny.

A comprehensive regression tree to estimate body weight from the skeleton.

The purpose of this research is to estimate actual body weight (in particular obesity) from the human skeleton. Known individuals (total n=187) were studied from the Bass Collection from the University of Tennessee, Knoxville. This research combines bone density, cross-sectional geometry of the femur and skeletal pathologies. Bone mineral density was calculated for the proximal femur. Three-dimensional bone surface models were created from computed tomographic scans. Cross-sectional geometry of the femur was calculated at five locations along the diaphysis. The pathologies analyzed were heel spurs, diffuse idiopathic skeletal hyperostosis (DISH), and tibial osteoarthritis. The best regression tree model included only four variables. The first split to estimate body weight was the minimum moment of inertia (I(y) ) at the distal femur, the second was midshaft width, then anteroposterior thickness at the proximal slice and the final split was the degree of DISH (SD 17.1-31.0 kg). The ability to estimate body weight from the skeleton is one more useful tool for the osteologist.