STRUCTURE AND FUNCTION OF THE GROUP III CHAPERONINS, A UNIQUE CLADE OF PROTEIN FOLDING NANOMACHINES

The survival and descent of cells is universally dependent on maintaining their proteins in a properly folded condition. It is widely accepted that the information for the folding of the nascent polypeptide chain into a native protein is encrypted in the amino acid sequence, and the Nobel Laureate Christian Anfinsen was the first to demonstrate that a protein could spontaneously refold after complete unfolding. However, it became clear that the observed folding rates for many proteins were much slower than rates estimated in vivo. This led to the recognition of required protein-protein interactions that promote proper folding. A unique group of proteins, the molecular chaperones, are responsible for maintaining protein homeostasis during normal growth as well as stress conditions. Chaperonins (CPNs) are ubiquitous and essential chaperones. They form ATP-dependent, hollow complexes that encapsulate polypeptides in two back-to-back stacked multisubunit rings, facilitating protein folding through highly cooperative allosteric articulation. CPNs are usually classified into Group I and Group II. Here, I report the characterization of a novel CPN belonging to a third Group, recently discovered in bacteria. Group III CPNs have close phylogenetic association to the Group II CPNs found in Archaea and Eukarya, and may be a relic of the Last Common Ancestor of the CPN family. The gene encoding the Group III CPN from Carboxydothermus hydrogenoformans and Candidatus Desulforudis audaxviator was cloned in E. coli and overexpressed in order to both characterize the protein and to demonstrate its ability to function as an ATPase chaperone. The opening and closing cycle of the Chy chaperonin was examined via site-directed mutations affecting the ATP binding site at R155. To relate the mutational analysis to the structure of the CPN, the crystal structure of both the AMP-PNP (an ATP analogue) and ADP bound forms were obtained in collaboration with Sun-Shin Cha in Seoul, South Korea. The ADP and ATP binding site substitutions resulted in frozen forms of the structures in open and closed conformations. From this, mutants were designed to validate hypotheses regarding key ATP interacting sites as well as important stabilizing interactions, and to observe the physical properties of the resulting complexes by calorimetry.

An Examination of Parental Engagement During the Middle School Years

The purpose of this quantitative study was to explore the previously unexamined phenomenon of middle school parental engagement in a large urban/suburban/rural school district of 209 schools in the mid-Atlantic region of the United States. Across 22 middle schools serving grades six-eight, this study collected and examined perceptions of the three key adult stakeholder groups – administrators, teachers, and parents – most actively involved in middle school parental engagement as described within the theoretical framework of academic socialization. Their reports of observable parental engagement activities were used to document how district stakeholders operationalize behaviors that represent the five actionable constructs and three themes of academic socialization to determine how the district “fares” in employing academic socialization as a middle school parent engagement strategy. The study also applied quantitative descriptive analysis through a one-way ANOVA to determine the significance of observable variations in actionable constructs between the perspectives of the three stakeholder groups. Finally, the study illuminated, through regression modeling, when confounding factors/independent variables such as race, income, school size, administrator and teacher experience, parents’ educational background, etc., impacted operationalization of academic socialization behaviors for middle school parent and family engagement. Rejecting the null hypothesis, the study found that the three stakeholder groups had statistically significant differences in perceptions of their implementation of activities aligned to academic socialization. This study ultimately illuminated ways in which these adult stakeholder groups share similar and varied perceptions about their engagement actions that support the achievement and maturation of middle school students. Significantly, this study provided key findings that illuminated areas that can be systemically addressed to transform middle school parent engagement practices through applied academic socialization theory into consistent and effective collaborative efforts between the home and school. The process of operationalizing academic socialization was outlined in terms that any school or district can follow to improve programs and practices of middle school parental engagement to serve in the best interests of students during this period of great transition for both child/adolescent growth and development and adult navigation of systems to provide support for students in this unique stage of growth and maturation.