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Structural and Genetic Studies Demonstrate Neurologic Dysfunction in Triosephosphate Isomerase Deficiency Is Associated with Impaired Synaptic Vesicle Dynamics.

Roland BP, Zeccola AM, Larsen SB, Amrich CG, Talsma AD, Stuchul KA, Heroux A, Levitan ES, VanDemark AP, Palladino MJ - PLoS Genet. (2016)

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology & Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, United States of America.

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.

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hTPIΔcat models predict that hTPIΔcat::hTPIG76E heterodimerization alters the TPI dimer interface.(A) The R75 side chain may insert itself into the active site in the context of a hTPIΔcat::hTPIT75R heterodimer. Shown is a superposition of hTPIWT (grey), hTPIΔcat (blue), and hTPIΔcat::hTPIT75R (green) obtained from Rosetta Backrub modeling. Loop3, containing T75R and G76E, is indicated in tan. For clarity, the mainchain of hTPIΔcat::hTPIT75R has been omitted. (B) Repositioning of E104 and R98 side chains assists the dimer interface to accommodate the G76E substitution in Rosetta modeling. Superposition of hTPIΔcat structure (blue) with the hTPIΔcat::hTPIG76E model (green). In both structures, Loop 3 is shown in tan and selected active site pocket residues are indicated. (C) Modeling the effect of T75R and G76E substitutions in the context of hTPIWT and hTPIΔcat structures. Normalized scores for the best 10% of 50 simulations were averaged for each experimental structure with the indicated computational substitution. Higher scores indicate a resulting model that is less favorable using the Rosetta energy function.
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pgen.1005941.g009: hTPIΔcat models predict that hTPIΔcat::hTPIG76E heterodimerization alters the TPI dimer interface.(A) The R75 side chain may insert itself into the active site in the context of a hTPIΔcat::hTPIT75R heterodimer. Shown is a superposition of hTPIWT (grey), hTPIΔcat (blue), and hTPIΔcat::hTPIT75R (green) obtained from Rosetta Backrub modeling. Loop3, containing T75R and G76E, is indicated in tan. For clarity, the mainchain of hTPIΔcat::hTPIT75R has been omitted. (B) Repositioning of E104 and R98 side chains assists the dimer interface to accommodate the G76E substitution in Rosetta modeling. Superposition of hTPIΔcat structure (blue) with the hTPIΔcat::hTPIG76E model (green). In both structures, Loop 3 is shown in tan and selected active site pocket residues are indicated. (C) Modeling the effect of T75R and G76E substitutions in the context of hTPIWT and hTPIΔcat structures. Normalized scores for the best 10% of 50 simulations were averaged for each experimental structure with the indicated computational substitution. Higher scores indicate a resulting model that is less favorable using the Rosetta energy function.

Mentions: While the hTPIK13M substitution (hTPIΔcat) resulted in multiple rearrangements that mimic the open or non-catalytic TPI conformation, the dimeric interface remained essentially unchanged, including the peptide backbone of Loop3 and sidechain positions hT75, hG76, and hM82 (Fig 5). To address how hTPIT75R and hTPIG76E substitutions may influence the dimer interface, we generated models of heterodimers in which one subunit contained either hT75R, hG76E, or both hT75R and hG76E substitutions, while the other monomer remained unaltered. Models were made using either wild-type TPI hTPIWT (PDB: 4POC) or the hTPIΔcat (PDB: 4ZVJ) as the structural template, and subjected to analysis by RosettaBackrub [65]. Briefly, Rosetta scores are predictions of the most energetically stable conformations with higher scores indicating less favorable positioning of the model. The algorithm was run 50 times for each mutation to be modeled. Of these 50 simulations, the lowest scores of hTPIΔcat::hTPIT75R and hTPIΔcat::hTPIG76E were selected and shown (Fig 9A and 9B), while the modeled structures whose Rosetta scores fell within the best 10% of its respective ensemble were collected for analysis (Fig 9C).


Structural and Genetic Studies Demonstrate Neurologic Dysfunction in Triosephosphate Isomerase Deficiency Is Associated with Impaired Synaptic Vesicle Dynamics.

Roland BP, Zeccola AM, Larsen SB, Amrich CG, Talsma AD, Stuchul KA, Heroux A, Levitan ES, VanDemark AP, Palladino MJ - PLoS Genet. (2016)

hTPIΔcat models predict that hTPIΔcat::hTPIG76E heterodimerization alters the TPI dimer interface.(A) The R75 side chain may insert itself into the active site in the context of a hTPIΔcat::hTPIT75R heterodimer. Shown is a superposition of hTPIWT (grey), hTPIΔcat (blue), and hTPIΔcat::hTPIT75R (green) obtained from Rosetta Backrub modeling. Loop3, containing T75R and G76E, is indicated in tan. For clarity, the mainchain of hTPIΔcat::hTPIT75R has been omitted. (B) Repositioning of E104 and R98 side chains assists the dimer interface to accommodate the G76E substitution in Rosetta modeling. Superposition of hTPIΔcat structure (blue) with the hTPIΔcat::hTPIG76E model (green). In both structures, Loop 3 is shown in tan and selected active site pocket residues are indicated. (C) Modeling the effect of T75R and G76E substitutions in the context of hTPIWT and hTPIΔcat structures. Normalized scores for the best 10% of 50 simulations were averaged for each experimental structure with the indicated computational substitution. Higher scores indicate a resulting model that is less favorable using the Rosetta energy function.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4816394&req=5

pgen.1005941.g009: hTPIΔcat models predict that hTPIΔcat::hTPIG76E heterodimerization alters the TPI dimer interface.(A) The R75 side chain may insert itself into the active site in the context of a hTPIΔcat::hTPIT75R heterodimer. Shown is a superposition of hTPIWT (grey), hTPIΔcat (blue), and hTPIΔcat::hTPIT75R (green) obtained from Rosetta Backrub modeling. Loop3, containing T75R and G76E, is indicated in tan. For clarity, the mainchain of hTPIΔcat::hTPIT75R has been omitted. (B) Repositioning of E104 and R98 side chains assists the dimer interface to accommodate the G76E substitution in Rosetta modeling. Superposition of hTPIΔcat structure (blue) with the hTPIΔcat::hTPIG76E model (green). In both structures, Loop 3 is shown in tan and selected active site pocket residues are indicated. (C) Modeling the effect of T75R and G76E substitutions in the context of hTPIWT and hTPIΔcat structures. Normalized scores for the best 10% of 50 simulations were averaged for each experimental structure with the indicated computational substitution. Higher scores indicate a resulting model that is less favorable using the Rosetta energy function.
Mentions: While the hTPIK13M substitution (hTPIΔcat) resulted in multiple rearrangements that mimic the open or non-catalytic TPI conformation, the dimeric interface remained essentially unchanged, including the peptide backbone of Loop3 and sidechain positions hT75, hG76, and hM82 (Fig 5). To address how hTPIT75R and hTPIG76E substitutions may influence the dimer interface, we generated models of heterodimers in which one subunit contained either hT75R, hG76E, or both hT75R and hG76E substitutions, while the other monomer remained unaltered. Models were made using either wild-type TPI hTPIWT (PDB: 4POC) or the hTPIΔcat (PDB: 4ZVJ) as the structural template, and subjected to analysis by RosettaBackrub [65]. Briefly, Rosetta scores are predictions of the most energetically stable conformations with higher scores indicating less favorable positioning of the model. The algorithm was run 50 times for each mutation to be modeled. Of these 50 simulations, the lowest scores of hTPIΔcat::hTPIT75R and hTPIΔcat::hTPIG76E were selected and shown (Fig 9A and 9B), while the modeled structures whose Rosetta scores fell within the best 10% of its respective ensemble were collected for analysis (Fig 9C).

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology & Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, United States of America.

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.

Show MeSH
Related in: MedlinePlus