Targeting Membrane Environments: The Role of Oleate Hydratase (OhyA) in Lipid-Protein Interactions Across Microbial Communities

/ABSTRACT Membrane integrity and function in biological systems heavily rely on lipid-protein interactions, particularly the role of proteins in concentrating signaling molecules within cellular membranes. Oleate hydratase (OhyA) is a microbial enzyme crucial in modifying unsaturated fatty acids (UFA) within cell membranes, transforming them into anti-inflammatory hydroxy fatty acids (hFA). This enzymatic activity is pivotal for various physiological processes, including gut health maintenance and immune response modulation. Despite its importance, the molecular mechanisms underpinning OhyA's membrane interaction and catalytic function, especially in pathogenic and commensal bacteria, remain unclear. Our research aims to elucidate the fundamental aspects of OhyA biology, focusing on three areas: investigation of how OhyA acquires UFA substrates from the membrane, understanding how bacteria utilize OhyA and distribute hFA to support colonization and immune modulation, and exploring the catalytic mechanisms of OhyA variants lacking typical functional domains. Our multidisciplinary approach leverages advanced structural biology, lipidomics, and genetic techniques to uncover novel insights into microbial physiology and protein- membrane interactions. This innovative framework promises significant contributions to both fundamental science and applied biotechnology. Characterizing OhyA and its role in lipid metabolism and immune modulation is pivotal for understanding bacterial pathogenesis and commensalism. Insights gained will inform biotechnological advancements, potentially facilitating the development of novel therapeutic strategies targeting microbial lipid metabolism. Our recent advances include solving crystal structures of OhyA•liposome complexes and identifying membrane- targeting domains, developing robust lipidomics pipelines for hFA quantification in biological samples, and establishing PPAR alpha as a key target for immunomodulatory hFA signaling. Our future investigations will expand on these foundations: defining the ecological relevance of OhyA in intestinal microbiomes using diverse murine models, determining the molecular details of OhyA-membrane interactions across bacterial taxa, and elucidating catalytic mechanisms of unusual OhyA variants and their implications for enzyme engineering. In summary, our research program aims to advance understanding of OhyA-mediated lipid metabolism in microbial communities, paving the way for innovations in microbiome science and biotechnological applications. Project Number: 1R35GM159612-01 | Fiscal Year: 2025 | NIH Institute/Center: National Institute of General Medical Sciences (NIGMS) | Principal Investigator: Christopher Radka | Institution: UNIVERSITY OF KENTUCKY, LEXINGTON, KY | Award Amount: $406,652 | Activity Code: R35 | Study Section: Maximizing Investigators' Research Award B Study Section[MRAB] View on NIH RePORTER: https://reporter.nih.gov/project-details/11197323