Predicting the structural basis of targeted protein degradation by integrating molecular dynamics simulations with structural mass spectrometry.

Targeted protein degradation (TPD) is a promising approach in drug discovery for degrading proteins implicated in diseases. A key step in this process is the formation of a ternary complex where a heterobifunctional molecule induces proximity of an E3 ligase to a protein of interest (POI), thus facilitating ubiquitin transfer to the POI. In this work, we characterize 3 steps in the TPD process. (1) We simulate the ternary complex formation of SMARCA2 bromodomain and VHL E3 ligase by combining hydrogen-deuterium exchange mass spectrometry with weighted ensemble molecular dynamics (MD). (2) We characterize the conformational heterogeneity of the ternary complex using Hamiltonian replica exchange simulations and small-angle X-ray scattering. (3) We assess the ubiquitination of the POI in the context of the full Cullin-RING Ligase, confirming experimental ubiquitinomics results. Differences in degradation efficiency can be explained by the proximity of lysine residues on the POI relative to ubiquitin.

Evaluation of production parameters and health of dairy cows treated with pegbovigrastim in the transition period.

The aim of this study was to evaluate the effects of pegbovigrastim injection (Imrestor, Elanco Animal Health, Greenfield, IN) on production parameters and postpartum disease occurrence (retained placenta, metritis, displaced abomasum, and clinical mastitis) in dairy cows. Study cows (n = 270) were blocked by parity group (multiparous or primiparous) and randomly assigned to control (CON, n = 144) or pegbovigrastim treatment (IMR, n = 126). Ten ± 4 days before expected calving and again at calving, IMR cows received 2.7 mL of Imrestor and CON cows received 2.7 mL of 0.9 % saline. Milk yield, fat, protein, lactose, solids nonfat (SNF) percent, and somatic cell count (SCC), body condition, hygiene, and lameness were evaluated weekly. Animals were evaluated for metritis twice weekly through rectal temperature, palpation, and uterine discharge evaluation until 30 days in milk. Farm personnel recorded other postpartum diseases. The MIXED procedure of SAS was used to evaluate milk composition and milk yields were analyzed as repeated measures in time with block, treatment, calving month, and lactation week included in mixed models. The GLIMMIX procedure was used to evaluate mastitis and metritis occurrence. Variables entered a model if P ≤ 0.10 when screened individually. Variables with P ≤ 0.10 were kept in the final model. Milk yield, fat, protein, lactose, SNF, and log of SCC were not significantly affected by treatment. Clinical mastitis occurrence did not differ between treatments, but only 17 cases were detected. Compared to CON, IMR treated cows had increased clinical metritis incidence, and were 2.46 times more likely to develop the disease. No difference was observed on puerperal metritis ocurrence. Cows given pegbovigrastim injections had increased odds of developing clinical signs of metritis, but no negative effects on milk production or composition were observed.