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Antivirulence strategy has been explored as an alternative to traditional antibiotic development. The bacterial type IV pilus is a virulence factor involved in host invasion and colonization in many antibiotic resistant pathogens. The PilB ATPase hydrolyzes ATP to drive the assembly of the pilus filament from pilin subunits. We evaluated Chloracidobacterium thermophilum PilB (CtPilB) as a model for structure-based virtual screening by molecular docking and molecular dynamics (MD) simulations. A hexameric structure of CtPilB was generated through homology modeling based on an existing crystal structure of a PilB from Geobacter metallireducens. Four representative structures were obtained from molecular dynamics simulations to examine the conformational plasticity of PilB and improve docking analyses by ensemble docking. Structural analyses after 1 μs of simulation revealed conformational changes in individual PilB subunits are dependent on ligand presence. Further, ensemble virtual screening of a library of 4234 compounds retrieved from the ZINC15 database identified five promising PilB inhibitors. Molecular docking and binding analyses using the four representative structures from MD simulations revealed that top-ranked compounds interact with multiple Walker A residues, one Asp-box residue, and one arginine finger, indicating these are key residues in inhibitor binding within the ATP binding pocket. The use of multiple conformations in molecular screening can provide greater insight into compound flexibility within receptor sites and better inform future drug development for therapeutics targeting the type IV pilus assembly ATPase. 

ABSTRACT Undergraduate research experiences (UREs) cultivate workforce skills, such as critical thinking, project management, and scientific communication. Many UREs in biophysical research have constraints related to limited resources, often resulting in smaller student cohorts, barriers for students entering a research environment, and fewer mentorship opportunities for graduate students. In response to those limitations, we have created a structured URE model that uses an asynchronous training style paired with direct-tiered mentoring delivered by peers, graduate students, and faculty. The adaptive undergraduate research training and experience (AURTE) framework was piloted as part of the Brown Experiential Learning program, a computational biophysics research lab. The program previously demonstrated substantial increases and improvements in the number of students served and skills developed. Here, we discuss the long-term effectiveness of the framework, impacts on graduate and undergraduate students, and efficacy in teaching research skills and computational-based biophysical methods. The longitudinal impact of our structured URE on student outcomes was analyzed by using student exit surveys, interviews, assessments, and 5 years of feedback from alumni. Results indicate high levels of student retention in research compared with university-wide metrics. Also, student feedback emphasizes how tiered mentoring enhanced research skill retention, while allowing graduate mentors to develop mentorship and workforce skills to expedite research. Responses from alumni affirm that workforce-ready skills (communicating science, data management, and scientific writing) acquired in the program persisted and were used in postgraduate careers. The framework reinforces the importance of establishing, iterating, and evaluating a structured URE framework to foster student success in biophysical research, while promoting mentorship skill training for graduate students. Future work will explore the adaptability of the framework in wet lab environments and probe the potential of AURTE in broader educational contexts. 

Abstract Chemotaxis is a fundamental process whereby bacteria seek out nutrient sources and avoid harmful chemicals. For the symbiotic soil bacteriumSinorhizobium meliloti, the chemotaxis system also plays an essential role in the interaction with its legume host. The chemotactic signaling cascade is initiated through interactions of an attractant or repellent compound with chemoreceptors or methyl‐accepting chemotaxis proteins (MCPs).S. melilotipossesses eight chemoreceptors to mediate chemotaxis. Six of these receptors are transmembrane proteins with periplasmic ligand‐binding domains (LBDs). The specific functions of McpW and McpZ are still unknown. Here, we report the crystal structure of the periplasmic domain of McpZ (McpZPD) at 2.7 Å resolution. McpZPD assumes a novel fold consisting of three concatenated four‐helix bundle modules. Through phylogenetic analyses, we discovered that this helical tri‐modular domain fold arose within the Rhizobiaceae family and is still evolving rapidly. The structure, offering a rare view of a ligand‐free dimeric MCP‐LBD, reveals a novel dimerization interface. Molecular dynamics calculations suggest ligand binding will induce conformational changes that result in large horizontal helix movements within the membrane‐proximal domains of the McpZPD dimer that are accompanied by a 5 Å vertical shift of the terminal helix toward the inner cell membrane. These results suggest a mechanism of transmembrane signaling for this family of MCPs that entails both piston‐type and scissoring movements. The predicted movements terminate in a conformation that closely mirrors those observed in related ligand‐bound MCP‐LBDs.