Mutations that cause threonine sensitivity identify catalytic and regulatory regions of the aspartate kinase of Saccharomyces cerevisiae

M Arévalo-Rodríguez; I L Calderón; S Holmberg

doi:10.1002/(SICI)1097-0061(19990930)15:13<1331::AID-YEA460>3.0.CO;2-W

Mutations that cause threonine sensitivity identify catalytic and regulatory regions of the aspartate kinase of Saccharomyces cerevisiae

M Arévalo-Rodríguez, I L Calderón, S Holmberg

Functional genomics

Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › peer review

16 Citationer (Scopus)

Abstract

Udgivelsesdato: 1999-Sep-30

Originalsprog	Engelsk
Tidsskrift	Yeast
Vol/bind	15
Udgave nummer	13
Sider (fra-til)	1331-45
Antal sider	14
ISSN	0749-503X
DOI	https://doi.org/10.1002/(SICI)1097-0061(19990930)15:13<1331::AID-YEA460>3.0.CO;2-W
Status	Udgivet - 1999

Bibliografisk note

Keywords: Alleles; Allosteric Regulation; Amino Acid Sequence; Aspartate Kinase; Catalysis; Molecular Sequence Data; Mutation; Saccharomyces cerevisiae; Structure-Activity Relationship; Threonine

Adgang til dokumentet

10.1002/(SICI)1097-0061(19990930)15:13<1331::AID-YEA460>3.0.CO;2-W

Citationsformater

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title = "Mutations that cause threonine sensitivity identify catalytic and regulatory regions of the aspartate kinase of Saccharomyces cerevisiae",

abstract = "The HOM3 gene of Saccharomyces cerevisiae encodes aspartate kinase, which catalyses the first step in the branched pathway leading to the synthesis of threonine and methionine from aspartate. Regulation of the carbon flow into this pathway takes place mainly by feedback inhibition of this enzyme by threonine. We have isolated and characterized three HOM3 mutants that show growth inhibition by threonine due to a severe, threonine-induced reduction of the carbon flow into the aspartate pathway, leading to methionine limitation. One of the mutants has an aspartate kinase which is 30-fold more strongly inhibited by threonine than the wild-type enzyme. The predicted amino acid substitution in this mutant, A406T, is located in a region associated with the modulation of the enzymatic activity. The other two mutants carry an aspartate kinase with reduced affinity for its substrates, aspartate and ATP. The corresponding amino acid substitutions, K26I and G25D, affect residues located in the vicinity of a highly conserved lysine-phenylalanine-glycine-glycine (KFGG) stretch present in the N-terminal part of the aspartate kinase, to which no function has so far been assigned. We suggest that this region is involved in substrate binding. Mutagenesis of a HOM3 region centred in the KFGG-coding triplets generated alleles that determine threonine sensitivity or auxotrophy for threonine and methionine, but not a phenotype associated with a feedback-resistant aspartate kinase, indicating that this region is not involved in the allosteric response of the enzyme.",

author = "M Ar{\'e}valo-Rodr{\'i}guez and Calder{\'o}n, {I L} and S Holmberg",

note = "Keywords: Alleles; Allosteric Regulation; Amino Acid Sequence; Aspartate Kinase; Catalysis; Molecular Sequence Data; Mutation; Saccharomyces cerevisiae; Structure-Activity Relationship; Threonine",

year = "1999",

doi = "10.1002/(SICI)1097-0061(19990930)15:13<1331::AID-YEA460>3.0.CO;2-W",

language = "English",

volume = "15",

pages = "1331--45",

journal = "Yeast",

issn = "0749-503X",

publisher = "JohnWiley & Sons Ltd",

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TY - JOUR

T1 - Mutations that cause threonine sensitivity identify catalytic and regulatory regions of the aspartate kinase of Saccharomyces cerevisiae

AU - Arévalo-Rodríguez, M

AU - Calderón, I L

AU - Holmberg, S

N1 - Keywords: Alleles; Allosteric Regulation; Amino Acid Sequence; Aspartate Kinase; Catalysis; Molecular Sequence Data; Mutation; Saccharomyces cerevisiae; Structure-Activity Relationship; Threonine

PY - 1999

Y1 - 1999

N2 - The HOM3 gene of Saccharomyces cerevisiae encodes aspartate kinase, which catalyses the first step in the branched pathway leading to the synthesis of threonine and methionine from aspartate. Regulation of the carbon flow into this pathway takes place mainly by feedback inhibition of this enzyme by threonine. We have isolated and characterized three HOM3 mutants that show growth inhibition by threonine due to a severe, threonine-induced reduction of the carbon flow into the aspartate pathway, leading to methionine limitation. One of the mutants has an aspartate kinase which is 30-fold more strongly inhibited by threonine than the wild-type enzyme. The predicted amino acid substitution in this mutant, A406T, is located in a region associated with the modulation of the enzymatic activity. The other two mutants carry an aspartate kinase with reduced affinity for its substrates, aspartate and ATP. The corresponding amino acid substitutions, K26I and G25D, affect residues located in the vicinity of a highly conserved lysine-phenylalanine-glycine-glycine (KFGG) stretch present in the N-terminal part of the aspartate kinase, to which no function has so far been assigned. We suggest that this region is involved in substrate binding. Mutagenesis of a HOM3 region centred in the KFGG-coding triplets generated alleles that determine threonine sensitivity or auxotrophy for threonine and methionine, but not a phenotype associated with a feedback-resistant aspartate kinase, indicating that this region is not involved in the allosteric response of the enzyme.

AB - The HOM3 gene of Saccharomyces cerevisiae encodes aspartate kinase, which catalyses the first step in the branched pathway leading to the synthesis of threonine and methionine from aspartate. Regulation of the carbon flow into this pathway takes place mainly by feedback inhibition of this enzyme by threonine. We have isolated and characterized three HOM3 mutants that show growth inhibition by threonine due to a severe, threonine-induced reduction of the carbon flow into the aspartate pathway, leading to methionine limitation. One of the mutants has an aspartate kinase which is 30-fold more strongly inhibited by threonine than the wild-type enzyme. The predicted amino acid substitution in this mutant, A406T, is located in a region associated with the modulation of the enzymatic activity. The other two mutants carry an aspartate kinase with reduced affinity for its substrates, aspartate and ATP. The corresponding amino acid substitutions, K26I and G25D, affect residues located in the vicinity of a highly conserved lysine-phenylalanine-glycine-glycine (KFGG) stretch present in the N-terminal part of the aspartate kinase, to which no function has so far been assigned. We suggest that this region is involved in substrate binding. Mutagenesis of a HOM3 region centred in the KFGG-coding triplets generated alleles that determine threonine sensitivity or auxotrophy for threonine and methionine, but not a phenotype associated with a feedback-resistant aspartate kinase, indicating that this region is not involved in the allosteric response of the enzyme.

U2 - 10.1002/(SICI)1097-0061(19990930)15:13<1331::AID-YEA460>3.0.CO;2-W

DO - 10.1002/(SICI)1097-0061(19990930)15:13<1331::AID-YEA460>3.0.CO;2-W

M3 - Journal article

C2 - 10509015

VL - 15

SP - 1331

EP - 1345

JO - Yeast

JF - Yeast

SN - 0749-503X

IS - 13

ER -