MOLE MECH OF THE LEGIONELLA PNEUMOPHILA EFFECTOR PROTEIN VIPF IN THE

Cornell University, Ithaca NY

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Abstract

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Legionella pneumophila is a facultative gram negative bacterium that can infect humans and cause a severe pneumonia known as Legionnaires[unreadable][unreadable]" disease. The bacteria are phagocytozed by macrophages and translocate effector proteins through a Type IV secretion system into the host cytoplasm. These effector proteins facilitate the survival and replication of the bacteria in a vacuole like compartment through several different mechanisms. VipF (Vacuolar protein sorting Inhibitor Protein F) was identified by a pathogen effector screen to identify Legionella effector proteins that are able to alter host cell protein trafficking pathways. Analysis of the VipF primary sequence reveals two GCN5-related acetyltransferase (GNAT) domains. Homology modeling along with small-angle X-ray scattering (SAXS), performed at CHESS, suggest that VipF is a bilobal monomeric protein. GNATs are a very diverse superfamily of proteins that are responsible for catalyzing the transfer of an acetyl group from acetyl-CoA to a primary amine of an acceptor peptide or small molecule. There are many reports of protein acetylation in diverse biological processes but little is known about what enzymes are responsible for each modification event and what its role is in regulating membrane trafficking. This research proposal aims to study the enzymatic activity of VipF and the mechanism by which it may alter host trafficking factors through a combination of genetics and cell biology coupled with biochemistry and structural biology. The major goals are to determine the molecular structure of VipF, elucidate its enzymatic mechanism and acetylated target, and understand how the enzymatic activity alters the vacuole protein sorting pathway in yeast. This work is of fundamental importance to understand how the bacterial effector protein VipF is able aid Legionella to form a permissive vacuole that undergo normal phagolysosome maturation.