Literature summary for 1.4.1.2 extracted from

Grzechowiak, M.; Sliwiak, J.; Jaskolski, M.; Ruszkowski, M.
Structural studies of glutamate dehydrogenase (isoform 1) from Arabidopsis thaliana, an important enzyme at the branch-point between carbon and nitrogen metabolism (2020), Front. Plant Sci., 11, 754 .

Application

Application	Comment	Organism
drug development	although AtGDH1 is insensitive to MPD in activity assays, several (+/-)-2-methyl-2,4-pentanediol (MPD) binding sites with conserved sequence are identified and the observation of druggable sites opens a potential for non-competitive herbicide design	Arabidopsis thaliana

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant AtGDH1 in the apo-form and in complex with NAD+, X-ray diffraction structure determination and analysis at 2.59 and 2.03 A resolution, respectively. Most of the subunits in the crystal structures, including those in NAD+ complex, are in open conformation, with domain II forming a large (albeit variable) angle with domain I. One of the subunits of the AtGDH1-NAD+ hexamer contains a serendipitous 2-oxoglutarate molecule in the active site, causing a dramatic closure of the domains	Arabidopsis thaliana
structure of recombinant GDH1 in the apo-form and in complex with NAD+ at 2.59 and 2.03 A resolution, respectively. Both in the apo form and in 1:1 complex with NAD+, it forms D3-symmetric homohexamers. A subunit of GDH1 consists of domain I, which is involved in hexamer formation and substrate binding, and of domain II which binds coenzyme	Arabidopsis thaliana

Crystallization (Comment)

Organism

purified recombinant AtGDH1 in the apo-form and in complex with NAD+, X-ray diffraction structure determination and analysis at 2.59 and 2.03 A resolution, respectively. Most of the subunits in the crystal structures, including those in NAD+ complex, are in open conformation, with domain II forming a large (albeit variable) angle with domain I. One of the subunits of the AtGDH1-NAD+ hexamer contains a serendipitous 2-oxoglutarate molecule in the active site, causing a dramatic closure of the domains

Arabidopsis thaliana

structure of recombinant GDH1 in the apo-form and in complex with NAD+ at 2.59 and 2.03 A resolution, respectively. Both in the apo form and in 1:1 complex with NAD+, it forms D3-symmetric homohexamers. A subunit of GDH1 consists of domain I, which is involved in hexamer formation and substrate binding, and of domain II which binds coenzyme

Arabidopsis thaliana

Inhibitors

Inhibitors	Comment	Organism
2-methyl-2,4-pentanediol	8.5 mM, 94% residual activity. the sequence contains several binding sites for 2-methyl-2,4-pentanediol	Arabidopsis thaliana
Ca2+	0.1 mM, 82% residual activity; 8% inhibition at 0.1 mM, 23% at 1 mM	Arabidopsis thaliana
Co2+	0.1 mM, 77% residual activity; 23% inhibition at 0.1 mM	Arabidopsis thaliana
Cu2+	0.1 mM, 65% residual activity; 35% inhibition at 0.1 mM	Arabidopsis thaliana
additional information	not inhibitory: Mn2+	Arabidopsis thaliana
Zn2+	0.1 mM, 51% residual activity; 49% inhibition at 0.1 mM	Arabidopsis thaliana

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
2.5	5	L-glutamate	pH 7.5, 25°C	Arabidopsis thaliana
2.5	5	L-glutamate	with NAD+, recombinant enzyme, pH and temperature not specified in the publication	Arabidopsis thaliana

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
mitochondrion	the N-terminal peptide preceding domain I is a mitochondrial targeting signal, the predicted cleavage site is Leu17-Leu18 followed by an potassium coordination site (Ser27, Ile30)	Arabidopsis thaliana	5739	-
mitochondrion	the N-terminal peptide preceding domain I is a mitochondrial targeting signal, with a predicted cleavage site for mitochondrial processing peptidase (MPP) at Leu17-Leu18 that is followed by an unexpected potassium coordination site (Ser27, Ile30)	Arabidopsis thaliana	5739	-

Metals/Ions

Metals/Ions	Comment	Organism	Structure
additional information	GDH1 shows no sensitivity to Mn2+	Arabidopsis thaliana

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
L-glutamate + H2O + NAD+	Arabidopsis thaliana	AtGDH1 activity in the forward reaction (oxidative deamination) is physiologically more relevant due to the high NAD+/NADH ratio in plant mitochondria	2-oxoglutarate + NH3 + NADH + H+	-	r

Organism

Organism	UniProt	Comment	Textmining
Arabidopsis thaliana	Q43314	-	-

Posttranslational Modification

Posttranslational Modification	Comment	Organism
proteolytic modification	the N-terminal peptide preceding domain I is a mitochondrial targeting signal, the predicted cleavage site is Leu17-Leu18 followed by an potassium coordination site (Ser27, Ile30)	Arabidopsis thaliana
proteolytic modification	the N-terminal peptide preceding domain I is a mitochondrial targeting signal, with a predicted cleavage site for mitochondrial processing peptidase (MPP) at Leu17-Leu18 that is followed by an unexpected potassium coordination site (Ser27, Ile30)	Arabidopsis thaliana

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
L-glutamate + H2O + NAD+	-	Arabidopsis thaliana	2-oxoglutarate + NH3 + NADH + H+	-	?
L-glutamate + H2O + NAD+	-	Arabidopsis thaliana	2-oxoglutarate + NH3 + NADH + H+	-	r
L-glutamate + H2O + NAD+	AtGDH1 activity in the forward reaction (oxidative deamination) is physiologically more relevant due to the high NAD+/NADH ratio in plant mitochondria	Arabidopsis thaliana	2-oxoglutarate + NH3 + NADH + H+	-	r

Subunits

Subunits	Comment	Organism
homohexamer	both in the apo form and in 1:1 complex with NAD+, it forms D3-symmetric homohexamers, composed of alpha-subunits	Arabidopsis thaliana
More	one subunit of AtGDH1 consists of domain I, which is involved in hexamer formation and substrate binding, and domain II, which binds the coenzyme. Most of the subunits in the crystal structures, including those in NAD+ complex, are in open conformation, with domain II forming a large (albeit variable) angle with domain I. One of the subunits of the AtGDH1-NAD+ hexamer contains a serendipitous 2-oxoglutarate molecule in the active site, causing a dramatic closure of the domains	Arabidopsis thaliana

Synonyms

Synonyms	Comment	Organism
At5g18170	locus name	Arabidopsis thaliana
AtGDH1	-	Arabidopsis thaliana
GDH1	-	Arabidopsis thaliana
glutamate dehydrogenase isoform 1	-	Arabidopsis thaliana
type I GDH	-	Arabidopsis thaliana

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism	Structure
13.3	-	L-glutamate	pH 7.5, 25°C	Arabidopsis thaliana
13.3	-	L-glutamate	with NAD+, recombinant enzyme, pH and temperature not specified in the publication	Arabidopsis thaliana

Cofactor

Cofactor	Comment	Organism
additional information	no detectable activity with NADP+	Arabidopsis thaliana
additional information	AtGDH1 in the presence of NADP+ instead of NAD+ shows no activity. Dynamics of the coafctor binding domain II, cofactor binding mode, overview	Arabidopsis thaliana
NAD+	-	Arabidopsis thaliana
NAD+	one NAD+ binds molecule per AtGDH1 subunit, Kd value 0.072 mM	Arabidopsis thaliana
NADH	-	Arabidopsis thaliana

General Information

General Information	Comment	Organism
evolution	GDHs are members of a superfamily of ELFV (Glu/Leu/Phe/Val) amino acid dehydrogenases and are subdivided into three subclasses, based on coenzyme specificity: NAD+-specific, NAD+/NADP+ dual-specific, and NADP+-specific. The mitochondrial AtGDH1 isozyme from Arabidopsis thaliana is NAD+-specific. Arabidopsis thaliana expresses three GDH isozymes (AtGDH1-3) targeted to mitochondria, of which AtGDH2 has an extra EF-hand motif and is stimulated by calcium, while AtGDH1's sensitivity to calcium is negligible. In vivo the AtGDH1-3 enzymes form homo- and heterohexamers of varied composition. Phylogenetic analysis of GDHs in plants. Plants have distinct isozymes of GDH that are either NAD or NADP-specific. NAD-specific GDHs are localized in mitochondria, whereas NADP-specific GDHs exist in chloroplasts. The sequence region 257-264 in AtGDH1 and AtGDH2, which directly precedes the EF-hand motif in AtGDH2 (residues 265-277), is the most altered region of AtGDH2 in comparison with AtGDH1	Arabidopsis thaliana
metabolism	GDH contributes to Glu homeostasis and plays a significant role at the junction of carbon and nitrogen assimilation pathways	Arabidopsis thaliana
additional information	several (+/-)-2-methyl-2,4-pentanediol (MPD) binding sites with conserved sequence are identified, but AtGDH1 is insensitive to MPD in activity assays. Structure function analysis of AtGDH1, overview. The open-to-closed conformational transition is required to form a fully functional active site	Arabidopsis thaliana
physiological function	glutamate dehydrogenase (GDH) releases ammonia in a reversible NAD(P)+-dependent oxidative deamination of glutamate that yields 2-oxoglutarate (2OG). Plants have distinct isozymes of GDH that are either NAD or NADP-specific. NAD-specific GDHs are localized in mitochondria, whereas NADP-specific GDHs exist in chloroplasts	Arabidopsis thaliana

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism	Structure
5.22	-	L-glutamate	with NAD+, recombinant enzyme, pH and temperature not specified in the publication	Arabidopsis thaliana