Ammonium transport proteins with changes in one of the conserved pore histidines have different performance in ammonia and methylamine conduction

Jinan Wang, Tim Fulford, Qiang Shao, Arnaud Javelle, Huaiyu Yang, Weiliang Zhu, Mike Merrick

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Two conserved histidine residues are located near the mid-point of the conduction channel of ammonium transport proteins. The role of these histidines in ammonia and methylamine transport was evaluated by using a combination of in vivo studies, molecular dynamics (MD) simulation, and potential of mean force (PMF) calculations. Our in vivo results showed that a single change of either of the conserved histidines to alanine leads to the failure to transport methylamine but still facilitates good growth on ammonia, whereas double histidine variants completely lose their ability to transport both methylamine and ammonia. Molecular dynamics simulations indicated the molecular basis of the in vivo observations. They clearly showed that a single histidine variant (H168A or H318A) of AmtB confines the rather hydrophobic methylamine more strongly than ammonia around the mutated sites, resulting in dysfunction in conducting the former but not the latter molecule. PMF calculations further revealed that the single histidine variants form a potential energy well of up to 6 kcal/mol for methylamine, impairing conduction of this substrate. Unlike the single histidine variants, the double histidine variant, H168A/H318A, of AmtB was found to lose its unidirectional property of transporting both ammonia and methylamine. This could be attributed to a greatly increased frequency of opening of the entrance gate formed by F215 and F107, in this variant compared to wild-type, with a resultant lowering of the energy barrier for substrate to return to the periplasm.

LanguageEnglish
Article numbere62745
Number of pages12
JournalPLOS One
Volume8
Issue number5
DOIs
Publication statusPublished - 7 May 2013

Fingerprint

methylamine
transport proteins
Ammonium Compounds
Ammonia
histidine
Histidine
Carrier Proteins
ammonia
molecular dynamics
Molecular Dynamics Simulation
Molecular dynamics
Periplasm
Energy barriers
energy
Computer simulation
Substrates
in vivo studies
Potential energy
Alanine
alanine

Keywords

  • ammonia
  • biological transport
  • cation transport proteins
  • conserved sequence
  • escherichia coli proteins
  • histidine
  • methylamines
  • molecular dynamics simulation
  • mutagenesis
  • mutation
  • protein conformation
  • saccharomyces cerevisiae proteins

Cite this

Wang, Jinan ; Fulford, Tim ; Shao, Qiang ; Javelle, Arnaud ; Yang, Huaiyu ; Zhu, Weiliang ; Merrick, Mike. / Ammonium transport proteins with changes in one of the conserved pore histidines have different performance in ammonia and methylamine conduction. In: PLOS One. 2013 ; Vol. 8, No. 5.
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Ammonium transport proteins with changes in one of the conserved pore histidines have different performance in ammonia and methylamine conduction. / Wang, Jinan; Fulford, Tim; Shao, Qiang; Javelle, Arnaud; Yang, Huaiyu; Zhu, Weiliang; Merrick, Mike.

In: PLOS One, Vol. 8, No. 5, e62745, 07.05.2013.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Ammonium transport proteins with changes in one of the conserved pore histidines have different performance in ammonia and methylamine conduction

AU - Wang, Jinan

AU - Fulford, Tim

AU - Shao, Qiang

AU - Javelle, Arnaud

AU - Yang, Huaiyu

AU - Zhu, Weiliang

AU - Merrick, Mike

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N2 - Two conserved histidine residues are located near the mid-point of the conduction channel of ammonium transport proteins. The role of these histidines in ammonia and methylamine transport was evaluated by using a combination of in vivo studies, molecular dynamics (MD) simulation, and potential of mean force (PMF) calculations. Our in vivo results showed that a single change of either of the conserved histidines to alanine leads to the failure to transport methylamine but still facilitates good growth on ammonia, whereas double histidine variants completely lose their ability to transport both methylamine and ammonia. Molecular dynamics simulations indicated the molecular basis of the in vivo observations. They clearly showed that a single histidine variant (H168A or H318A) of AmtB confines the rather hydrophobic methylamine more strongly than ammonia around the mutated sites, resulting in dysfunction in conducting the former but not the latter molecule. PMF calculations further revealed that the single histidine variants form a potential energy well of up to 6 kcal/mol for methylamine, impairing conduction of this substrate. Unlike the single histidine variants, the double histidine variant, H168A/H318A, of AmtB was found to lose its unidirectional property of transporting both ammonia and methylamine. This could be attributed to a greatly increased frequency of opening of the entrance gate formed by F215 and F107, in this variant compared to wild-type, with a resultant lowering of the energy barrier for substrate to return to the periplasm.

AB - Two conserved histidine residues are located near the mid-point of the conduction channel of ammonium transport proteins. The role of these histidines in ammonia and methylamine transport was evaluated by using a combination of in vivo studies, molecular dynamics (MD) simulation, and potential of mean force (PMF) calculations. Our in vivo results showed that a single change of either of the conserved histidines to alanine leads to the failure to transport methylamine but still facilitates good growth on ammonia, whereas double histidine variants completely lose their ability to transport both methylamine and ammonia. Molecular dynamics simulations indicated the molecular basis of the in vivo observations. They clearly showed that a single histidine variant (H168A or H318A) of AmtB confines the rather hydrophobic methylamine more strongly than ammonia around the mutated sites, resulting in dysfunction in conducting the former but not the latter molecule. PMF calculations further revealed that the single histidine variants form a potential energy well of up to 6 kcal/mol for methylamine, impairing conduction of this substrate. Unlike the single histidine variants, the double histidine variant, H168A/H318A, of AmtB was found to lose its unidirectional property of transporting both ammonia and methylamine. This could be attributed to a greatly increased frequency of opening of the entrance gate formed by F215 and F107, in this variant compared to wild-type, with a resultant lowering of the energy barrier for substrate to return to the periplasm.

KW - ammonia

KW - biological transport

KW - cation transport proteins

KW - conserved sequence

KW - escherichia coli proteins

KW - histidine

KW - methylamines

KW - molecular dynamics simulation

KW - mutagenesis

KW - mutation

KW - protein conformation

KW - saccharomyces cerevisiae proteins

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DO - 10.1371/journal.pone.0062745

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JO - PLOS One

T2 - PLOS One

JF - PLOS One

SN - 1932-6203

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M1 - e62745

ER -