Pyruvate phosphate dikinase: sequence of the histidyl peptide, the pyrophosphoryl and phosphoryl carrier

Goss, Neil H.; Evans, Claudia T.; Wood, Harland G.

doi:10.1021/bi00566a022

Cited by 39 publications

(29 citation statements)

References 23 publications

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“…The purification protocol for R. rubrum PPDK described here resembles that for the C4 leaf enzyme (25) and those for the B. symbiosus (10,11) and A. aceti (23) places the DEAE-cellulose step before (NH4)2SO4 fractionation because this sequence markedly decreases the time and number of steps involved; the HSSF is immediately mixed with DEAE-cellulose without prior desalting, and the 0.2 M KCl batch eluant is then immediately fractionated in a single step and stored for up to 1 month at -80°C as a 0 to 55% (NH4)2SO4 precipitate. Unlike the above protocols, which end with molecular sieve chromatography, ours terminates with a rapid negative purification step with Blue A, which yields a 98% recovery and a three-to fourfold increase in specific activity.…”

Section: Discussionmentioning

confidence: 99%

Partial purification and characterization of pyruvate, orthophosphate dikinase from Rhodospirillum rubrum

Ernst¹,

Budde²,

Chollet³

1986

J Bacteriol

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We confirmed an earlier report (B. B. Buchanan, J. Bacteriol. 119:1066Bacteriol. 119: -1068Bacteriol. 119: , 1974) that the nonsulfur purple photosynthetic bacterium Rhodospirillum rubrum contains pyruvate,orthophosphate dikinase (EC 2.7.9.1) activity that is absolutely dependent upon all three substrates by performing enzyme assays in both the forward (phosphoenolpyruvate formation) and reverse (ATP formation) directions. Of the various carbon sources tested, photoheterotrophic growth on DL-lactate plus bicarbonate proved to be best for the production of dikinase activity units. A four-step protocol, which included batch DEAE-cellulose processing, ammonium sulfate fractionation, and chromatography on hydroxylapatite and Blue A Dyematrex gels, was devised for partially purifying the enzyme from such cells. The protein was purified about 80-fold to an apparent electrophoretic purity of about 60% and a final specific activity of 3.6 U/mg of protein, with about a 35% overall recovery of activity units. Estimations of native and monomeric relative molecular weights by sucrose density gradient centrifugation, high-pressure liquid chromatography-based size exclusion chromatography, denaturing electrophoresis, and immunoblotting suggested that the holoenzyme was most likely a homodimer of 92.7-kilodalton subunits. The results are compared with related previous data on the nonphotosynthetic bacterial dikinase and the C4 mesophyll chloroplast enzyme.

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Section: Discussionmentioning

confidence: 99%

Partial purification and characterization of pyruvate, orthophosphate dikinase from Rhodospirillum rubrum

Ernst¹,

Budde²,

Chollet³

1986

J Bacteriol

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“…Enzyme phosphorylation occurs on the N8 atom of a histidine residue (4-6), which has been identified as His-455 in the amino acid sequence of PPDK from Clostridium symbiosum, the subject of this study (6 …”

Section: Introductionmentioning

confidence: 99%

Swiveling-domain mechanism for enzymatic phosphotransfer between remote reaction sites.

Herzberg

Chen

Kapadia

et al. 1996

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

144

205

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“…3) is the site of Enzyme I phosphorylation by HPr(His-P) (Meadow et al, 1990) and of PPDK by ATP or phosphoenolpyruvate (PEP) (Goss et al, 1980;Burnell, 1984;Roeske et al, 1988). The preceding threonyl residue (threoninezl5 in the aligned proteins shown in Fig.…”

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confidence: 99%

Sequence analyses and evolutionary relationships among the energy‐coupling proteins enzyme I and HPr of the bacterial phosphoenolpyruvate: Sugar phosphotransferase system

et al. 1993

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We have previously reported the overexpression, purification, and biochemical properties of the Bacillus subtilis Enzyme I of the phosphoeno1pyruvate:sugar phosphotransferase system (PTS) (Reizer, J., et al., 1992, J. Biol. Chem. 267,9158-9169). We now report the sequencing of thepfsl gene of B. subtilis encoding Enzyme I (570 amino acids and 63,076 Da). Putative transcriptional regulatory signals are identified, and the p f s operon is shown to be subject to carbon source-dependent regulation. Multiple alignments of the B. subtilis Enzyme I with (1) six other sequenced Enzymes I of the PTS from various bacterial species, (2) phosphoenolpyruvate synthase of Escherichia coli, and (3) bacterial and plant pyruvate:phosphate dikinases (PPDKs) revealed regions of sequence similarity as well as divergence. Statistical analyses revealed that these three types of proteins comprise a homologous family, and the phylogenetic tree of the 1 1 sequenced protein members of this family was constructed. This tree was compared with that of the 12 sequenced HPr proteins or protein domains. Antibodies raised against the B. subtilis and E. coli Enzymes I exhibited immunological cross-reactivity with each other as well as with PPDK of Bacferoides syrnbiosus, providing support for the evolutionary relationships of these proteins suggested from the sequence comparisons. Putative flexible linkers tethering the N-terminal and the C-terminal domains of protein members of the Enzyme I family were identified, and their potential significance with regard to Enzyme I function is discussed. The codon choice pattern of the B. subtilis and E, coliptsl andptsH genes was found to exhibit a bias toward optimal codons in these organisms. Quantitative analysis of this bias indicated that theptsHgenes have higher bias toward codons favored in highly expressed genes than the p t s l genes, in agreement with their relative levels of expression.

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Pyruvate phosphate dikinase: sequence of the histidyl peptide, the pyrophosphoryl and phosphoryl carrier

Cited by 39 publications

References 23 publications

Partial purification and characterization of pyruvate, orthophosphate dikinase from Rhodospirillum rubrum

Partial purification and characterization of pyruvate, orthophosphate dikinase from Rhodospirillum rubrum

Swiveling-domain mechanism for enzymatic phosphotransfer between remote reaction sites.

Sequence analyses and evolutionary relationships among the energy‐coupling proteins enzyme I and HPr of the bacterial phosphoenolpyruvate: Sugar phosphotransferase system

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