Journal Article

Caspase-4 disaggregates lipopolysaccharide micelles via LPS-CARD interaction

Seong-Ho Kim, Kyung Eun Lee, Jinsu An, Min Jung Kim, Hak Suk Chung, Eun Gyeong Yang, Dohyeon Hwang, So Yeon Kim

2019

Overview

Lipopolysaccharides (LPS) are a major component of the outer membrane of Gram-negative bacteria and are pathogen-associated molecular patterns recognized by the TLR4/MD2 complex that induces an inflammatory response. Recently, the cytosolic receptors caspase-4/-5/-11 that bind LPS inside the cell and trigger inflammasome activation or pyroptosis, have been identified. Despite the important roles of caspase-4 in human immune responses, few studies have investigated its biochemical characteristics and interactions with LPS. Since caspase-4 (C258A) purified from an Escherichia coli host forms aggregates, monomeric proteins including full-length caspase-4, caspase-4 (C258A), and the CARD domain of caspase-4 have been purified from the insect cell system. Here, we report the overexpression and purification of monomeric caspase-4 (C258A) and CARD domain from E . coli and demonstrate that purified caspase-4 (C258A) and CARD domain bind large LPS micelles and disaggregate them to small complexes. As the molar ratio of caspase-4 to LPS increases, the size of the caspase-4/LPS complex decreases. Our results present a new function of caspase-4 and set the stage for structural and biochemical studies, and drug discovery targeting LPS/caspase-4 interactions by establishing a facile purification method to obtain large quantities of purified caspase-4 (C258A) and the CARD domain.

• Publisher: Springer Science and Business Media LLC,Nature Publishing Group UK,Nature Publishing Group
• Subject: 631/250/256/2177 / 631/45/287/1190 / 82 / 82/80 / 82/83 / Bacteria / Bacterial Outer Membrane / Biochemical characteristics / Biochemistry / Caspase / Caspase Activation and Recruitment Domain / Caspase-4 / Caspases, Initiator / Catalytic Domain / Cell activation / Cell Line / Chromatography / Drug discovery / E coli / Enzymes / Escherichia coli / Gram-negative bacteria / HEK293 Cells / Humanities and Social Sciences / Humans / Immune response / Inflammasomes / Inflammation / Lipids / Lipopolysaccharides / Micelles / Molecular weight / multidisciplinary / Pathogens / Protein Binding / Proteins / Pyroptosis / Roles / Science / Science (multidisciplinary) / TLR4 protein / Toll-like receptors