Preparation of Crystalline Nucleoside Diphosphate Kinase from Baker's Yeast and Identification of 1‐[<sup>32</sup>P]Phosphohistidine as the Main Phosphorylated Product of an Alkaline Hydrolysate of Enzyme Incubated with Adenosine [<sup>32</sup>P]Triphosphate

Edlund, Bror; Rask, Lars; Olsson, Pelle; Wålinder, Olov; Zetterqvist, Örjan; Engström, Lorentz

doi:10.1111/j.1432-1033.1969.tb00630.x

Cited by 48 publications

(24 citation statements)

References 19 publications

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“…1-Phosphohistidine has been found to occur in phosphate transfer reactions catalyzed by nucleotide diphosphate kinase (36) and Hpr, the phosphoenolpyruvate-dependent phosphocarrier protein of Escherichia coli (9) and Staphylococcus aureus (37).…”

Section: Discussionmentioning

confidence: 99%

Isolation of 3-phosphohistidine from phosphorylated pyruvate, phosphate dikinase.

Spronk¹,

Yoshida²,

Wood³

1976

Proc. Natl. Acad. Sci. U.S.A.

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(3,4) have demonstrated that the pyruvate,phosphate dikinase reaction occurs by a three-site nonclassical Tri (Uni Uni) Ping Pong mechanism. A pyrophosphoryl-and phosphoryl-histidyl residue of the enzyme has been proposed to serve as a carrier between the three sites (4).A number of enzymes that have reaction mechanisms involving phosphate moieties covalently bound to enzyme are now known (5). The phosphate has been found linked to proteins through different types of linkage, including a phosphoester with the hydroxyl group of serine (6), a phosphoramidate to N-1 or N-3 of histidine (7-10), and an acid anhydride to a y-carboxyl group of glutamic acid (11).The phosphoramidate and acid anhydride type of bond may be considered high in chemical energy (12), and thus either of these might meet the requirement of phosphoryl pyruvate, phosphate dikinase. However, at the outset we could not exclude the possibility of ester linkage to serine because the phosphate of this type of bond has been known to be transferable to ADP with concomitant formation of ATP (13). Thus, in an appropriate micro-environment even a seryl-phosphate bond may be considered a "high energy" type.In the present work using pyruvate,phosphate dikinase from Bacteroides symbiosus, the type of covalent bond of phosphoryl enzyme has been investigated and it has been shown to be a N-3-phosphohistidyl residue of the enzyme. (14). The product was purified by chromatography on a Dowex 1-X8 column (Cl-, 2 X 25 cm) (1). The radioactive fractions were pooled, lyophilized, and stored frozen. The concentration of P-enolpyruvate was determined enzymatically by a pyruvate kinase-lactate dehydrogenase system and specific radioactivity was calculated in cpm/,mol by dividing the cpm in material that migrated on paper chromatography with authentic P-enol[2-'4C]pyruvate by the Amols of P-enolpyruvate in the aliquot. The product was 92-94% pure. MATERIALS AND METHODS ChemicalsSynthesis of Phosphohistidine. Phosphohistidine was synthesized by reacting potassium phosphoramidate and histidine (15). Phosphoramidate was prepared from diphenylchlorophosphate (16). The phosphohistidine was purified by chromatography on a 2.4 X 30 cm column of Dowex 1-X8 using a 2-liter linear gradient of 0-1.0 M KHCO3, pH 8.25. The fractions were assayed for histidine (17) and phosphate (18), following cleavage of the phosphoramidate by acidification of an aliquot to pH 1.0. The fractions were pooled, cooled in ice, and brought to pH 3.0 with perchloric acid. After CO2 evolution ceased, the pH was rapidly adjusted to 9.0 with potassium hydroxide, and the sample was filtered to remove the precipitated potassium perchlorate and lyophilized. Upon rechromatography the product (with some potassium perchlorate) gave a single peak, which was 3-phosphohistidine, as confirmed by chromatography of the product with a tracer amount of a mixture of 1-and 3-phospho[carboxyl-'4C]histidine that had been prepared according to Hays et al. (10).Purification and Assay of Pyruvate,Phosphate Dikinase. The enzyme ...

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Section: Discussionmentioning

confidence: 99%

Isolation of 3-phosphohistidine from phosphorylated pyruvate, phosphate dikinase.

Spronk¹,

Yoshida²,

Wood³

1976

Proc. Natl. Acad. Sci. U.S.A.

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“…The NDP kinase reaction occurs via a ping-pong mechanism utilizing a phosphorylated enzyme intermediate, and the phosphorylation occurs on a histidine residue (15). We hypothesized that the multiple bands detected on isoelectric focusing gels represent phosphorylated intermediates of the hexameric AWD/NDP kinase, the most acidic isoform representing a hexamer with all six subunits phosphorylated at the active site.…”

Section: Purification Of Recombinant Awd/ndp Kinase-awd/ndpmentioning

confidence: 99%

“…The ␥-phosphate of a nucleoside triphosphate is first transferred to the active site histidine of the enzyme (15), forming an enzyme intermediate. The ␥-phosphate is then transferred from the enzyme to a nucleoside diphosphate acceptor.…”

mentioning

confidence: 99%

Point Mutations in awd Which Revert the Prune/Killer of Prune Lethal Interaction Affect Conserved Residues That Are Involved in Nucleoside Diphosphate Kinase Substrate Binding and Catalysis

Timmons

Hersperger

et al. 1995

Journal of Biological Chemistry

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The awd gene of Drosophila melanogaster encodes a nucleoside diphosphate kinase. Killer of prune (Kpn) is a mutation in the awd gene which substitutes Ser for Pro at position 97 and causes dominant lethality in individuals that do not have a functional prune gene. This lethality is not due to an inadequate amount of nucleoside diphosphate (NDP) kinase activity. In order to understand why the prune/Killer of prune combination is lethal, even in the presence of an adequate NDP kinase specific activity level, and to understand the biochemical basis for the conditional lethality of the awd Kpn mutation, we generated second site mutations which revert this lethal interaction. All of the 12 revertants we recovered are second site mutations of the awd Kpn gene. Three revertants have deletions of the awd Kpn protein coding region. Two revertants have substitutions of the initiator methionine and do not accumulate KPN protein. Seven revertants have amino acid substitutions of conserved residues that are likely to affect the active site: five of these have no enzymatic activity and two have a very low level of specific activity. These data suggest that an altered NDP kinase activity is involved in the mechanism underlying the conditional lethality of the awd Kpn mutation.

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“…dominating i n an a l k a l i n e h y d r o l y s a t e o f phosphorylated NDP k i n a s e f r o m baker's y e a s t (3,4).…”

Section: N T E R M E D I a T E L Y Phosphorylated By I T S S U B S mentioning

confidence: 99%

Effect of Chemical Modification of a Histidine and a Lysine Residue of Pea Seed Nucleoside Diphosphate Kinase

Edlund

Heldin

Engström

1982

Upsala Journal of Medical Sciences

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Chemical m o d i f i c a t i o n o f a h i s t i d i n e and l y s i n e r e s i d u e i n a c t i v a t e s pea seed n u c l e o s i d e diphosphate k i n a s e (NDP k i n a s e ) . r e a c t i v e l y s i n e r e s i d u e , a t t h e a c t i v e s i t e o f pea seed NDP kinase, i n a d d it i o n t o t h e h i s t i d i n e r e s i d u e phosphorylated b y t h e s u b s t r a t e ATP as a con-sequence o f t h e enzyme r e a c t i o n . The presence o f a r e a c t i v e l y s i n e a t t h e

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Preparation of Crystalline Nucleoside Diphosphate Kinase from Baker's Yeast and Identification of 1‐[³²P]Phosphohistidine as the Main Phosphorylated Product of an Alkaline Hydrolysate of Enzyme Incubated with Adenosine [³²P]Triphosphate

Cited by 48 publications

References 19 publications

Isolation of 3-phosphohistidine from phosphorylated pyruvate, phosphate dikinase.

Isolation of 3-phosphohistidine from phosphorylated pyruvate, phosphate dikinase.

Point Mutations in awd Which Revert the Prune/Killer of Prune Lethal Interaction Affect Conserved Residues That Are Involved in Nucleoside Diphosphate Kinase Substrate Binding and Catalysis

Effect of Chemical Modification of a Histidine and a Lysine Residue of Pea Seed Nucleoside Diphosphate Kinase

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Preparation of Crystalline Nucleoside Diphosphate Kinase from Baker's Yeast and Identification of 1‐[32P]Phosphohistidine as the Main Phosphorylated Product of an Alkaline Hydrolysate of Enzyme Incubated with Adenosine [32P]Triphosphate

Cited by 48 publications

References 19 publications

Isolation of 3-phosphohistidine from phosphorylated pyruvate, phosphate dikinase.

Isolation of 3-phosphohistidine from phosphorylated pyruvate, phosphate dikinase.

Point Mutations in awd Which Revert the Prune/Killer of Prune Lethal Interaction Affect Conserved Residues That Are Involved in Nucleoside Diphosphate Kinase Substrate Binding and Catalysis

Effect of Chemical Modification of a Histidine and a Lysine Residue of Pea Seed Nucleoside Diphosphate Kinase

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Preparation of Crystalline Nucleoside Diphosphate Kinase from Baker's Yeast and Identification of 1‐[³²P]Phosphohistidine as the Main Phosphorylated Product of an Alkaline Hydrolysate of Enzyme Incubated with Adenosine [³²P]Triphosphate