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Information on EC 2.7.2.11 - glutamate 5-kinase and Organism(s) Saccharomyces cerevisiae and UniProt Accession P32264
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2.7.2.11 glutamate 5-kinaseIUBMB Comments
In the absence of downstream enzymes, the product rapidly cyclizes to 5-oxo-L-proline and phosphate.
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Word Map
- 2.7.2.11
- proba
- 1-pyrroline-5-carboxylate
- l-proline
-
osmotolerance
- delta1-pyrroline-5-carboxylate
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osmoprotection
- gpr
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catharanthus
-
uprooted
- biotechnology
The taxonomic range for the selected organisms is: Saccharomyces cerevisiae
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Synonyms
gamma-glutamyl kinase, glutamate 5-kinase, glutamate kinase, gamma-gk, glutamate-5-kinase, gamma-glutamate kinase, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
ATP-L-glutamate 5-phosphotransferase
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ATP:gamma-L-glutamate phosphotransferase
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gamma-glutamate kinase
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gamma-glutamyl kinase
gamma-glutamylphosphate kinase
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glutamate kinase
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GPK
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kinase (phosphorylating), glutamate
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kinase, glutamate (phosphorylating)
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gamma-glutamyl kinase
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
phospho group transfer
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PATHWAY SOURCE
PATHWAYS
KEGG
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-, -, -, -
SYSTEMATIC NAME
IUBMB Comments
ATP:L-glutamate 5-phosphotransferase
In the absence of downstream enzymes, the product rapidly cyclizes to 5-oxo-L-proline and phosphate.
CAS REGISTRY NUMBER
COMMENTARY
54596-30-4
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SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
ATP + L-glutamate
ADP + L-glutamate 5-phosphate
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-
-
?
ATP + L-glutamate
ADP + L-glutamate 5-phosphate
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-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
ATP + L-glutamate
ADP + L-glutamate 5-phosphate
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-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
L-proline
GK activity is subject to feedback inhibition by proline
additional information
no significant differences in growth between mutant yeast cells on SD agar medium with heat shock (50°C, 3 to 5 h), ethanol (16 to20%), and sorbitol (2.0 to 2.5 M) stresses, although the proline-accumulating cells are more resistant to these stresses than cells expressing wild-type GK
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additional information
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no significant differences in growth between mutant yeast cells on SD agar medium with heat shock (50°C, 3 to 5 h), ethanol (16 to20%), and sorbitol (2.0 to 2.5 M) stresses, although the proline-accumulating cells are more resistant to these stresses than cells expressing wild-type GK
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
GK activity requires gamma-glutamyl phosphate reductase
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additional information
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GK activity requires gamma-glutamyl phosphate reductase
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IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
14.3
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
malfunction
metabolism
physiological function
the enzyme has an alternative function independent of proline biosynthesis. Gene PRO1 genetically interacts with UBP3, which encodes ubiquitin-specific protease, and is required for selective autophagy of ribosomes (ribophagy). The enzyme activity is indispensable for ribophagy, ribophagy is important for cell survival during nitrogen starvation
additional information
malfunction
the DELTApro1 cells are more sensitive to various stresses than wild-type cells. Yeast cells with gene PRO1 deletion or expressing inactive enzyme display a defect for ribophagy but not for nonselective autophagy. Yeast strains deficient ribophagy are sensitive to nitrogen starvation
malfunction
Yeast cells expressing mutant enzymes D154N and I150T accumulate proline and show a higher tolerance to various stresses, including freezing, ethanol, osmotic pressure, and heat shock
metabolism
gamma-glutamyl kinase is a key enzyme in the Saccharomyces cerevisiae proline biosynthesis pathway
additional information
conserved residue Asp156 plays an important role in the catalytic activity
additional information
residues Asp154 and Ile150 in the Saccharomyces cerevisiae gamma-glutamate kinase are important for allosteric regulation and the affinity with L-proline and L-glutamate, but not with ATP. The PUA domain itself is required for the full display of enzyme activity, but is not involved in the allosteric regulation and the affinity with substrates and proline. The proline-auxotrophic yeast strain BY4741DELTApro1DELTAcar2 that expresses the PUA domain deletion-mutant grows even on proline-free medium, although a growth defect is observed
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
D154N
E149K
very insensitive to feedback inhibition, shows prominent increase in the proline content, freeze tolerance like the D154N mutant
I150T
N142D/I166V
very insensitive to feedback inhibition, shows prominent increase in the proline content, is more tolerant to freezing stress than cells expressing the D154N mutant
Q79H
the mutation results in extreme desensitization to feedback inhibition by L-proline, leading to L-proline overproduction. The relative activity of the variant is 96% in the presence of 100 mM L-proline and 87% in the presence of 400 mM L-proline. The specific activity of the variant is slightly reduced compared with the wild type enzyme
additional information
D154N
the mutation results in a prominant increase in cell viability after freezing at -20°C compared to the viability of the cells harboring the wild-type PRO1 gene. The altered gamma-glutamyl kinase results in stabilization of the complex with gamma-glutamyl phosphate reductase or has an indirect effect on gamma-glutamyl phosphate reductase activity which leads to an increase in L-proline production in Saccharomyces cerevisiae
D154N
is 64fold less sensitive to feedback inhibition than wild-type GK, leading to proline accumulation
D154N
site-directed mutagenesis, the mutant enzyme is less sensitive to proline feedback inhibition compared to the wild-type enzyme and shows an increased thermostability
I150T
very insensitive to feedback inhibition, shows prominent increase in the proline content, is more tolerant to freezing stress than cells expressing the D154N mutant
I150T
site-directed mutagenesis, the mutant enzyme is less sensitive to proline feedback inhibition compared to the wild-type enzyme and shows an increased thermostability
I150T
the mutant is greatly insensitive to feedback inhibition, leading to L-proline overproduction. The relative activity of the variant is 59% in the presence of 100 mM L-proline. The specific activity of the variant is slightly reduced compared with the wild type enzyme
additional information
construction of a gene PRO1 disruption mutant, phenotype, detailed overview. Overexpression of wild-type enzyme partially suppresses the phenotype of the DELTApro1 strain, which is deficient in ribophagy
additional information
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truncated yeast gamma-glutamyl kinase proteins are engineered from which the C-terminal region is deleted. A complementation test in Escherichia coli and yeast and enzymatic analysis of recombinant proteins reveal that a 67-residue linker sequence between a 255-residue kinase domain and a 106-residue archaeosine transglycosylase (PUA) domain is essential for gamma-glutamyl kinase activity. 67 or more residues of the C-terminal region, not the PUA domain itself, are required for the full enzymatic activity
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene PRO1, cloning of mutants in Escherichia coli strain DH5alpha
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
biotechnology
the D154N mutation results in a prominant increase in cell viability after freezing at -20°C compared to the viability of the cells harboring the wild-type PRO1 gene, method for breeding novel freeze-tolerant yeast strains
additional information
additional information
GK is the rate-limiting enzyme in the proline biosynthesis
additional information
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GK is the rate-limiting enzyme in the proline biosynthesis
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Morita, Y.; Nakamori, S.; Takagi, H.
L-proline accumulation and freeze tolerance of Saccharomyces cerevisiae are caused by a mutation in the PRO1 gene encoding gamma-glutamyl kinase
Appl. Environ. Microbiol.
69
212-219
2003
Saccharomyces cerevisiae (P32264), Saccharomyces cerevisiae
Sekine, T.; Kawaguchi, A.; Hamano, Y.; Takagi, H.
Desensitization of feedback inhibition of the Saccharomyces cerevisiae gamma-glutamyl kinase enhances proline accumulation and freezing tolerance
Appl. Environ. Microbiol.
73
4011-4019
2007
Saccharomyces cerevisiae (P32264), Saccharomyces cerevisiae
Kaino, T.; Tasaka, Y.; Tatehashi, Y.; Takagi, H.
Functional analysis of the C-terminal region of gamma-glutamyl kinase of Saccharomyces cerevisiae
Biosci. Biotechnol. Biochem.
76
454-461
2012
Saccharomyces cerevisiae
Tatehashi, Y.; Watanabe, D.; Takagi, H.
gamma-Glutamyl kinase is involved in selective autophagy of ribosomes in Saccharomyces cerevisiae
FEBS Lett.
590
2906-2914
2016
Saccharomyces cerevisiae (P32264)
Tatehashi, Y.; Takagi, H.
Characterization of gamma-glutamyl kinase mutants from Saccharomyces cerevisiae
J. Biosci. Bioeng.
116
576-579
2013
Saccharomyces cerevisiae (P32264)
Murakami, N.; Kotaka, A.; Isogai, S.; Ashida, K.; Nishimura, A.; Matsumura, K.; Hata, Y.; Ishida, H.; Takagi, H.
Effects of a novel variant of the yeast gamma-glutamyl kinase Pro1 on its enzymatic activity and sake brewing
J. Ind. Microbiol. Biotechnol.
47
715-723
2020
Saccharomyces cerevisiae (P32264)
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