Structural studies show that the A178L mutation in the C-terminal hinge of the catalytic loop-6 of triosephosphate isomerase (TIM) induces a closed-like conformation in dimeric and monomeric TIM

Markus Alahuhta, Marco G. Casteleijn, Peter Neubauer, Rik K. Wierenga

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Abstract

The flexible catalytic loop, loop-6, of TIM has evolved to preferably be open in the unliganded state and to preferably be closed in the liganded state. The N-terminal and C-terminal hinges of this loop are important for its opening/closing mechanism. In this study, a small conserved C-terminal hinge residue, Ala178, has been mutated into a residue with a larger side chain, Leu178. This mutation has been made in the dimeric trypanosomal wild-type TIM (wtTIM) and in its mutated catalytically competent monomeric variant (ml1TIM). The variants are referred to as A178L and ml1A178L, respectively. Crystal structures have been determined of unliganded A178L (at 2.2 Å), liganded A178L (at 1.89 Å), unliganded ml1A178L (at 2.3 Å) and liganded ml1A178L (at 1.18 Å) using the transition-state analogue 2-phosphoglycolate as a ligand. Structural characterization of the two variants shows that this mutation favours the closed conformation of the C-terminal hinge region, even in the absence of ligand. In the structure of the unliganded A178L variant a range of new loop-6 conformations are observed, including subunits in which the tip of loop-6 is completely disordered. The catalytic efficiency of A178L is lower than that of wtTIM, which correlates with the structural differences between the apo forms of wtTIM and A178L, in particular the more disordered loop-6 in the structure of unliganded A178L. In the liganded structures of A178L and ml1A178L the structural differences induced by the mutation are minimal. Structural characterization of the ml1A178L variant highlights its structural plasticity.
Original languageEnglish
Pages (from-to)178-188
Number of pages11
JournalActa Crystallographica Section D: Biological Crystallography
Volume64
Issue number2
DOIs
Publication statusPublished - 16 Jan 2008
MoE publication typeA1 Journal article-refereed

Fingerprint

Triose-Phosphate Isomerase
hinges
Hinges
mutations
Conformations
Mutation
Ligands
Plasticity
Crystal structure
ligands
closing
plastic properties
analogs
crystal structure

Keywords

  • Hinge
  • Loop-6
  • Monomeric TIM
  • Structural plasticity

Cite this

@article{f6572957e7b14487bc8f842e0f22ee96,
title = "Structural studies show that the A178L mutation in the C-terminal hinge of the catalytic loop-6 of triosephosphate isomerase (TIM) induces a closed-like conformation in dimeric and monomeric TIM",
abstract = "The flexible catalytic loop, loop-6, of TIM has evolved to preferably be open in the unliganded state and to preferably be closed in the liganded state. The N-terminal and C-terminal hinges of this loop are important for its opening/closing mechanism. In this study, a small conserved C-terminal hinge residue, Ala178, has been mutated into a residue with a larger side chain, Leu178. This mutation has been made in the dimeric trypanosomal wild-type TIM (wtTIM) and in its mutated catalytically competent monomeric variant (ml1TIM). The variants are referred to as A178L and ml1A178L, respectively. Crystal structures have been determined of unliganded A178L (at 2.2 {\AA}), liganded A178L (at 1.89 {\AA}), unliganded ml1A178L (at 2.3 {\AA}) and liganded ml1A178L (at 1.18 {\AA}) using the transition-state analogue 2-phosphoglycolate as a ligand. Structural characterization of the two variants shows that this mutation favours the closed conformation of the C-terminal hinge region, even in the absence of ligand. In the structure of the unliganded A178L variant a range of new loop-6 conformations are observed, including subunits in which the tip of loop-6 is completely disordered. The catalytic efficiency of A178L is lower than that of wtTIM, which correlates with the structural differences between the apo forms of wtTIM and A178L, in particular the more disordered loop-6 in the structure of unliganded A178L. In the liganded structures of A178L and ml1A178L the structural differences induced by the mutation are minimal. Structural characterization of the ml1A178L variant highlights its structural plasticity.",
keywords = "Hinge, Loop-6, Monomeric TIM, Structural plasticity",
author = "Markus Alahuhta and Casteleijn, {Marco G.} and Peter Neubauer and Wierenga, {Rik K.}",
year = "2008",
month = "1",
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doi = "10.1107/S0907444907059021",
language = "English",
volume = "64",
pages = "178--188",
journal = "Acta Crystallographica Section D: Structural Biology",
issn = "2059-7983",
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T1 - Structural studies show that the A178L mutation in the C-terminal hinge of the catalytic loop-6 of triosephosphate isomerase (TIM) induces a closed-like conformation in dimeric and monomeric TIM

AU - Alahuhta, Markus

AU - Casteleijn, Marco G.

AU - Neubauer, Peter

AU - Wierenga, Rik K.

PY - 2008/1/16

Y1 - 2008/1/16

N2 - The flexible catalytic loop, loop-6, of TIM has evolved to preferably be open in the unliganded state and to preferably be closed in the liganded state. The N-terminal and C-terminal hinges of this loop are important for its opening/closing mechanism. In this study, a small conserved C-terminal hinge residue, Ala178, has been mutated into a residue with a larger side chain, Leu178. This mutation has been made in the dimeric trypanosomal wild-type TIM (wtTIM) and in its mutated catalytically competent monomeric variant (ml1TIM). The variants are referred to as A178L and ml1A178L, respectively. Crystal structures have been determined of unliganded A178L (at 2.2 Å), liganded A178L (at 1.89 Å), unliganded ml1A178L (at 2.3 Å) and liganded ml1A178L (at 1.18 Å) using the transition-state analogue 2-phosphoglycolate as a ligand. Structural characterization of the two variants shows that this mutation favours the closed conformation of the C-terminal hinge region, even in the absence of ligand. In the structure of the unliganded A178L variant a range of new loop-6 conformations are observed, including subunits in which the tip of loop-6 is completely disordered. The catalytic efficiency of A178L is lower than that of wtTIM, which correlates with the structural differences between the apo forms of wtTIM and A178L, in particular the more disordered loop-6 in the structure of unliganded A178L. In the liganded structures of A178L and ml1A178L the structural differences induced by the mutation are minimal. Structural characterization of the ml1A178L variant highlights its structural plasticity.

AB - The flexible catalytic loop, loop-6, of TIM has evolved to preferably be open in the unliganded state and to preferably be closed in the liganded state. The N-terminal and C-terminal hinges of this loop are important for its opening/closing mechanism. In this study, a small conserved C-terminal hinge residue, Ala178, has been mutated into a residue with a larger side chain, Leu178. This mutation has been made in the dimeric trypanosomal wild-type TIM (wtTIM) and in its mutated catalytically competent monomeric variant (ml1TIM). The variants are referred to as A178L and ml1A178L, respectively. Crystal structures have been determined of unliganded A178L (at 2.2 Å), liganded A178L (at 1.89 Å), unliganded ml1A178L (at 2.3 Å) and liganded ml1A178L (at 1.18 Å) using the transition-state analogue 2-phosphoglycolate as a ligand. Structural characterization of the two variants shows that this mutation favours the closed conformation of the C-terminal hinge region, even in the absence of ligand. In the structure of the unliganded A178L variant a range of new loop-6 conformations are observed, including subunits in which the tip of loop-6 is completely disordered. The catalytic efficiency of A178L is lower than that of wtTIM, which correlates with the structural differences between the apo forms of wtTIM and A178L, in particular the more disordered loop-6 in the structure of unliganded A178L. In the liganded structures of A178L and ml1A178L the structural differences induced by the mutation are minimal. Structural characterization of the ml1A178L variant highlights its structural plasticity.

KW - Hinge

KW - Loop-6

KW - Monomeric TIM

KW - Structural plasticity

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DO - 10.1107/S0907444907059021

M3 - Article

C2 - 18219118

AN - SCOPUS:38549151788

VL - 64

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EP - 188

JO - Acta Crystallographica Section D: Structural Biology

JF - Acta Crystallographica Section D: Structural Biology

SN - 2059-7983

IS - 2

ER -