An, Jinsu Kim, Seong Ho Hwang, Dohyeon Lee, Kyung Eun Kim, Min Jung Yang, Eun Gyeong Kim, So Yeon Chung, Hak Suk 2024-01-19T21:03:35Z 2024-01-19T21:03:35Z 2021-09-02 2019-01 2045-2322 https://pubs.kist.re.kr/handle/201004/120545 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. English Nature Publishing Group Caspase-4 disaggregates lipopolysaccharide micelles via LPS-CARD interaction Article 10.1038/s41598-018-36811-4 1 Scientific Reports, v.9 Scientific Reports 9 Y scie scopus 000456955500006 2-s2.0-85060828947 Multidisciplinary Sciences Science & Technology - Other Topics Article ESCHERICHIA-COLI BINDING RECOGNITION ACTIVATION PROTEINS Inflammatory caspase lipopolysaccharide caspase-4 LPS-caspase-4 interaction CARD domain