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1.
PLoS Genet ; 12(3): e1005941, 2016 Mar.
Article in English | MEDLINE | ID: mdl-27031109

ABSTRACT

Triosephosphate isomerase (TPI) deficiency is a poorly understood disease characterized by hemolytic anemia, cardiomyopathy, neurologic dysfunction, and early death. TPI deficiency is one of a group of diseases known as glycolytic enzymopathies, but is unique for its severe patient neuropathology and early mortality. The disease is caused by missense mutations and dysfunction in the glycolytic enzyme, TPI. Previous studies have detailed structural and catalytic changes elicited by disease-associated TPI substitutions, and samples of patient erythrocytes have yielded insight into patient hemolytic anemia; however, the neuropathophysiology of this disease remains a mystery. This study combines structural, biochemical, and genetic approaches to demonstrate that perturbations of the TPI dimer interface are sufficient to elicit TPI deficiency neuropathogenesis. The present study demonstrates that neurologic dysfunction resulting from TPI deficiency is characterized by synaptic vesicle dysfunction, and can be attenuated with catalytically inactive TPI. Collectively, our findings are the first to identify, to our knowledge, a functional synaptic defect in TPI deficiency derived from molecular changes in the TPI dimer interface.


Subject(s)
Anemia, Hemolytic, Congenital Nonspherocytic/genetics , Carbohydrate Metabolism, Inborn Errors/genetics , Drosophila melanogaster/genetics , Nervous System Diseases/genetics , Synaptic Vesicles/genetics , Triose-Phosphate Isomerase/deficiency , Triose-Phosphate Isomerase/genetics , Anemia, Hemolytic, Congenital Nonspherocytic/pathology , Animals , Behavior, Animal , Carbohydrate Metabolism, Inborn Errors/pathology , Crystallography, X-Ray , Dimerization , Humans , Mutation, Missense , Nervous System Diseases/pathology , Protein Conformation , Synaptic Vesicles/pathology , Triose-Phosphate Isomerase/chemistry , Triose-Phosphate Isomerase/metabolism
2.
Biochim Biophys Acta ; 1852(1): 61-9, 2015 Jan.
Article in English | MEDLINE | ID: mdl-25463631

ABSTRACT

Triosephosphate isomerase (TPI) is a glycolytic enzyme which homodimerizes for full catalytic activity. Mutations of the TPI gene elicit a disease known as TPI Deficiency, a glycolytic enzymopathy noted for its unique severity of neurological symptoms. Evidence suggests that TPI Deficiency pathogenesis may be due to conformational changes of the protein, likely affecting dimerization and protein stability. In this report, we genetically and physically characterize a human disease-associated TPI mutation caused by an I170V substitution. Human TPI(I170V) elicits behavioral abnormalities in Drosophila. An examination of hTPI(I170V) enzyme kinetics revealed this substitution reduced catalytic turnover, while assessments of thermal stability demonstrated an increase in enzyme stability. The crystal structure of the homodimeric I170V mutant reveals changes in the geometry of critical residues within the catalytic pocket. Collectively these data reveal new observations of the structural and kinetic determinants of TPI Deficiency pathology, providing new insights into disease pathogenesis.


Subject(s)
Anemia, Hemolytic, Congenital Nonspherocytic/pathology , Carbohydrate Metabolism, Inborn Errors/pathology , Catalytic Domain , Triose-Phosphate Isomerase/deficiency , Triose-Phosphate Isomerase/metabolism , Anemia, Hemolytic, Congenital Nonspherocytic/enzymology , Animals , Behavior, Animal , Carbohydrate Metabolism, Inborn Errors/enzymology , Disease Models, Animal , Drosophila , Enzyme Stability , Humans , Mutation , Triose-Phosphate Isomerase/chemistry , Triose-Phosphate Isomerase/genetics
3.
J Cell Sci ; 126(Pt 14): 3151-8, 2013 Jul 15.
Article in English | MEDLINE | ID: mdl-23641070

ABSTRACT

Triosephosphate isomerase (TPI) is a glycolytic enzyme that converts dihydroxyacetone phosphate (DHAP) into glyceraldehyde 3-phosphate (GAP). Glycolytic enzyme dysfunction leads to metabolic diseases collectively known as glycolytic enzymopathies. Of these enzymopathies, TPI deficiency is unique in the severity of neurological symptoms. The Drosophila sugarkill mutant closely models TPI deficiency and encodes a protein prematurely degraded by the proteasome. This led us to question whether enzyme catalytic activity was crucial to the pathogenesis of TPI sugarkill neurological phenotypes. To study TPI deficiency in vivo we developed a genomic engineering system for the TPI locus that enables the efficient generation of novel TPI genetic variants. Using this system we demonstrate that TPI sugarkill can be genetically complemented by TPI encoding a catalytically inactive enzyme. Furthermore, our results demonstrate a non-metabolic function for TPI, the loss of which contributes significantly to the neurological dysfunction in this animal model.


Subject(s)
Anemia, Hemolytic, Congenital Nonspherocytic/enzymology , Carbohydrate Metabolism, Inborn Errors/enzymology , Drosophila melanogaster/physiology , Longevity , Paralysis/enzymology , Triose-Phosphate Isomerase/deficiency , Triose-Phosphate Isomerase/metabolism , Anemia, Hemolytic, Congenital Nonspherocytic/genetics , Animals , Carbohydrate Metabolism, Inborn Errors/genetics , Catalysis , Dihydroxyacetone Phosphate/metabolism , Disease Models, Animal , Drosophila melanogaster/enzymology , Female , Gene Knockout Techniques , Genetic Complementation Test , Genetic Engineering , Glyceraldehyde 3-Phosphate/metabolism , Glycolysis/genetics , Hot Temperature/adverse effects , Male , Mutation/genetics , Paralysis/genetics , Stress, Physiological/genetics , Transgenes/genetics , Triose-Phosphate Isomerase/genetics
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