Structural and Biochemical Basis for the Inhibition of Cell Death by APIP, a Methionine Salvage Enzyme
소속 :
연사 : Prof. Jin Kuk Yang (Soongsil University)
일시 : 2014-04-03 16:30 ~
장소 : 25-1동 103호
일 시 : 2014년 4월 3일, 오후 4:30
장 소 : 25-1동 국제회의실
-Abstract-
PIP, Apaf-1 interacting protein, has been known to inhibit two main types of programmed cell death: apoptosis and pyroptosis. Seemingly related to this function, APIP was recently found to be associated with cancers and inflammatory diseases. Distinct from its inhibitory role in cell death, APIP was also shown to act as a 5-methylthioribulose-1-phosphate dehydratase, or MtnB, in the methionine salvage pathway. Here, we report the structural and enzymatic characterization of APIP as an MtnB enzyme with a Km of 9.32 μM and a Vmax of 1.39 μmol min-1 mg-1. The crystal structure was determined at 2.0 Å resolution, revealing an overall fold similar to members of the zinc-dependent class II aldolase family. APIP/MtnB exists as a tetramer in solution, and exhibits an assembly with C4 symmetry in the crystal lattice. The pocket-shaped active site harboring a zinc ion is located at the end of a long cleft between two adjacent subunits. We propose an enzymatic reaction mechanism involving Glu139* as a catalytic acid/base, as supported by enzymatic assays of active site mutants, substrate-docking study, and sequence conservation analysis. We explored the relationship between two distinct functions of APIP/MtnB, cell death inhibition and methionine salvage, by measuring the ability of enzymatic mutants to inhibit cell death, and determined that APIP/MtnB functions as a cell death inhibitor independently of its MtnB enzyme activity for apoptosis induced by either hypoxia or etoposide, but dependently for caspase-1-induced pyroptosis. Our results establish the structural and biochemical groundwork for future mechanistic studies of the role of APIP/MtnB in modulating cell death and inflammation, and the development of related diseases.
장 소 : 25-1동 국제회의실
-Abstract-
PIP, Apaf-1 interacting protein, has been known to inhibit two main types of programmed cell death: apoptosis and pyroptosis. Seemingly related to this function, APIP was recently found to be associated with cancers and inflammatory diseases. Distinct from its inhibitory role in cell death, APIP was also shown to act as a 5-methylthioribulose-1-phosphate dehydratase, or MtnB, in the methionine salvage pathway. Here, we report the structural and enzymatic characterization of APIP as an MtnB enzyme with a Km of 9.32 μM and a Vmax of 1.39 μmol min-1 mg-1. The crystal structure was determined at 2.0 Å resolution, revealing an overall fold similar to members of the zinc-dependent class II aldolase family. APIP/MtnB exists as a tetramer in solution, and exhibits an assembly with C4 symmetry in the crystal lattice. The pocket-shaped active site harboring a zinc ion is located at the end of a long cleft between two adjacent subunits. We propose an enzymatic reaction mechanism involving Glu139* as a catalytic acid/base, as supported by enzymatic assays of active site mutants, substrate-docking study, and sequence conservation analysis. We explored the relationship between two distinct functions of APIP/MtnB, cell death inhibition and methionine salvage, by measuring the ability of enzymatic mutants to inhibit cell death, and determined that APIP/MtnB functions as a cell death inhibitor independently of its MtnB enzyme activity for apoptosis induced by either hypoxia or etoposide, but dependently for caspase-1-induced pyroptosis. Our results establish the structural and biochemical groundwork for future mechanistic studies of the role of APIP/MtnB in modulating cell death and inflammation, and the development of related diseases.
