Journal: The Journal of Biological Chemistry

Article Title: IACS-010759, a potent inhibitor of glycolysis-deficient hypoxic tumor cells, inhibits mitochondrial respiratory complex I through a unique mechanism

doi: 10.1074/jbc.RA120.013366

Figure Lengend Snippet: Effects of IACS-010759 on alkynylation of 49 kDa-Asp160 by AL1. Alkynylation of 49 kDa-Asp160 in SMPs (2.0 mg of proteins/ml) was performed via LDT chemistry using AL1 (0.10 μm) in the presence of different concentrations of IACS-010759. The alkynylation can be visualized by conjugating a fluorescent tag, TAMRA-N3, to Asp160(COO)-(CH2)2-CCH via click chemistry after solubilizing SMPs. Bullatacin (10 μm) and BAY 87-2243 (30 μm) were used as references. Approximately 30 μg of proteins were loaded in each well. Top, gel image of SDS-PAGE analysis used for LDT chemistry; bottom, the extent of suppression by test compounds. Values in graphs are means ± S.E. (error bars) (n = 3).

Article Snippet: Materials Bullatacin, piericidin A, and fenpyroximate were kindly provided by J. L. McLaughlin (Purdue University, West Lafayette, IN), S. Yoshida (RIKEN, Saitama, Japan), and Nihon Nohyaku Co., Ltd. (Tokyo, Japan), respectively.

Techniques: SDS Page

Journal: The Journal of Biological Chemistry

Article Title: IACS-010759, a potent inhibitor of glycolysis-deficient hypoxic tumor cells, inhibits mitochondrial respiratory complex I through a unique mechanism

doi: 10.1074/jbc.RA120.013366

Figure Lengend Snippet: Effects of IACS-010759 on the specific binding of [125I]AzQ to the 49-kDa subunit. The photoaffinity labeling of the 49-kDa subunit by [125I]AzQ (5.0 nm) was carried out in SMPs (2.0 mg of proteins/ml) in the presence of different concentrations of IACS-010759 (up to 20 μm, 4000-fold excess). Bullatacin (5.0 μm), aminoquinazoline (5.0 μm), and BAY 87-2243 (20 μm) were used as references. Approximately 30 μg of proteins were loaded in each well. Top, gel image of SDS-PAGE analysis used for the photoaffinity labeling; bottom, the extent of suppression by test compounds. Values in graphs are means ± S.E. (error bars) (n = 3). n.s., not significant compared with control (one-way ANOVA followed by Dunnett's test).

Article Snippet: Materials Bullatacin, piericidin A, and fenpyroximate were kindly provided by J. L. McLaughlin (Purdue University, West Lafayette, IN), S. Yoshida (RIKEN, Saitama, Japan), and Nihon Nohyaku Co., Ltd. (Tokyo, Japan), respectively.

Techniques: Binding Assay, Labeling, SDS Page, Control

Journal: The Journal of Biological Chemistry

Article Title: IACS-010759, a potent inhibitor of glycolysis-deficient hypoxic tumor cells, inhibits mitochondrial respiratory complex I through a unique mechanism

doi: 10.1074/jbc.RA120.013366

Figure Lengend Snippet: Effects of IACS-010759 on the specific binding of [125I]S1QEL1.1-PD1 to the ND1 subunit. The photoaffinity labeling of the ND1 subunit by [125I]S1QEL1.1-PD1 (5.0 nm) was carried out in SMPs (2.0 mg of proteins/ml) in the presence of different concentrations of IACS-010759 (up to 20 μm, 4000-fold excess), followed by the isolation of complex I by BN-PAGE and resolution of complex I subunits by SDS-PAGE. Bullatacin (5.0 μm) and BAY 87-2243 (20 μm) were used as references. Approximately 30 μg of proteins were loaded in each well. Top, gel image of SDS-PAGE analysis used for the photoaffinity labeling; bottom, the extent of suppression by test compounds. Values in graphs are means ± S.E. (error bars) (n = 3). n.s., not significant among the three (one-way ANOVA followed by Tukey's test).

Article Snippet: Materials Bullatacin, piericidin A, and fenpyroximate were kindly provided by J. L. McLaughlin (Purdue University, West Lafayette, IN), S. Yoshida (RIKEN, Saitama, Japan), and Nihon Nohyaku Co., Ltd. (Tokyo, Japan), respectively.

Techniques: Binding Assay, Labeling, Isolation, SDS Page

Journal: The Journal of Biological Chemistry

Article Title: IACS-010759, a potent inhibitor of glycolysis-deficient hypoxic tumor cells, inhibits mitochondrial respiratory complex I through a unique mechanism

doi: 10.1074/jbc.RA120.013366

Figure Lengend Snippet: Effects of inhibitors on the events that take place above the third loop connecting TMHs 5 and 6 in the ND1 subunit

Article Snippet: Materials Bullatacin, piericidin A, and fenpyroximate were kindly provided by J. L. McLaughlin (Purdue University, West Lafayette, IN), S. Yoshida (RIKEN, Saitama, Japan), and Nihon Nohyaku Co., Ltd. (Tokyo, Japan), respectively.

Techniques: Modification, Binding Assay, Inhibition