Transfer of 1,3-diphosphoglycerate between glyceraldehyde 3-phosphate dehydrogenase and 3-phosphoglycerate kinase via an enzyme-substrate-enzyme complex

Weber, Jon P.; Bernhard, Sidney A.

doi:10.1021/bi00260a042

Cited by 109 publications

(68 citation statements)

References 22 publications

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“…GAPDH was found to be distributed in muscle cytosol and the SR membrane, and ultrastructural localization studies have also revealed that GAPDH along with other glycolytic enzymes are bound to the cytoplasmic face of the SR membrane in skeletal muscle (33,38). A role for GAPDH together with 3-phosphoglycerate kinase (PGK) has been previously established in the production of ATP from GAP, NAD ϩ , P i , and ADP at the level of the SR membrane (19,20,39,40). In addition, a recent study demonstrated that synaptic vesicles were capable of generating local ATP via the membrane-associated GAPDH system, which could support the accumulation of the excitatory neurotransmitter glutamate even during significantly reduced global cellular ATP concentrations (41).…”

Section: Camentioning

confidence: 99%

The Muscle-specific Calmodulin-dependent Protein Kinase Assembles with the Glycolytic Enzyme Complex at the Sarcoplasmic Reticulum and Modulates the Activity of Glyceraldehyde-3-phosphate Dehydrogenase in a Ca2+/Calmodulin-dependent Manner

Singh

Salih

Leddy

et al. 2004

Journal of Biological Chemistry

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The skeletal muscle specific Ca 2؉ /calmodulin-dependent protein kinase (CaMKII␤ M ) is localized to the sarcoplasmic reticulum (SR) by an anchoring protein, ␣KAP, but its function remains to be defined. Protein interactions of CaMKII␤ M indicated that it exists in complex with enzymes involved in glycolysis at the SR membrane. The kinase was found to complex with glycogen phosphorylase, glycogen debranching enzyme, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and creatine kinase in the SR membrane. CaMKII␤ M was also found to assemble with aldolase A, GAPDH, enolase, lactate dehydrogenase, creatine kinase, pyruvate kinase, and phosphorylase b kinase from the cytosolic fraction. The interacting proteins were substrates of CaMKII␤ M , and their phosphorylation was enhanced in a Ca 2؉ -and calmodulin (CaM)-dependent manner. The CaMKII␤ M could directly phosphorylate GAPDH and markedly increase (ϳ3.4-fold) its activity in a Ca 2؉ /CaMdependent manner. These data suggest that the muscle CaMKII␤ M isoform may serve to assemble the glycogenmobilizing and glycolytic enzymes at the SR membrane and specifically modulate the activity of GAPDH in response to calcium signaling. Thus, the activation of CaMKII␤ M in response to calcium signaling would serve to modulate GAPDH and thereby ATP and NADH levels at the SR membrane, which in turn will regulate calcium transport processes.Free calcium (Ca 2ϩ ) regulates diverse cellular functions by acting as an intracellular second messenger. A large part of these cellular functions are mediated by CaM, 1 which is the ubiquitous intracellular Ca 2ϩ receptor. The Ca 2ϩ ⅐CaM complex allosterically activates numerous proteins, including Ca 2ϩ /CaMdependent protein kinase II (CaMKII) (1). CaMKII is a multifunctional enzyme that is highly expressed in brain and muscle. The kinase is believed to serve important roles in synaptic transmission (2, 3), gene transcription (4, 5), cell growth (6), and control of excitation-contraction coupling (7-9).The subcellular distribution of CaMKII indicates cytosolic and membrane localizations in different tissues (10). In skeletal muscle, an isoform of CaMKII is targeted to the SR membrane by a non-kinase protein, ␣KAP (11, 12). Different studies have been conducted to determine whether membrane-bound CaM kinase could phosphorylate different substrate proteins by virtue of its proximity effects and thereby regulate SR function. Although the calcium release channel/ryanodine receptor (RyR) and the calcium pump/Ca 2ϩ -ATPase in skeletal muscle SR were shown to be substrates of CaMKII␤ (13, 14), there does not appear to be any clear effects on the regulation of functional activity of these proteins induced by such phosphorylation (7,8,(15)(16)(17). Moreover, there is clear evidence that the RyR and calcium pump are regulated by local ATP, Ca 2ϩ , and CaM through direct ligand binding (15)(16)(17). In this regard, both Ca 2ϩ and CaM are present at the SR, and the level of ATP is believed to be tightly controlled through a membrane-bound glycolytic mac...

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Section: Camentioning

confidence: 99%

The Muscle-specific Calmodulin-dependent Protein Kinase Assembles with the Glycolytic Enzyme Complex at the Sarcoplasmic Reticulum and Modulates the Activity of Glyceraldehyde-3-phosphate Dehydrogenase in a Ca2+/Calmodulin-dependent Manner

Singh

Salih

Leddy

et al. 2004

Journal of Biological Chemistry

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“…The activities of GAPDH and 3-PGK are energetically coupled, utilizing GAP, NAD, P i , and ADP, to yield 3-PG, ATP, NADH, and H ϩ (46,47). Because GAPDH is enriched in synaptic vesicles, we considered the possibility that 3-PGK may also preferentially localize to synaptic vesicle membranes.…”

Section: A 37-kda Protein Enriched In Synaptic Vesicles Is Labeled Wimentioning

confidence: 99%

“…2). Bernhard and co-workers (46,47) have demonstrated direct transfer of 1,3-BPG from GAPDH to 3-PGK in muscle tissue. Inhibition of vesicle-bound GAPDH activity by iodoacetate led to a reduction of ATP synthesis as well as to an attenuation of Glu uptake into synaptic vesicles (Fig.…”

Section: Atp Produced By Glycolysis Is Predominantly Utilized For Vesmentioning

confidence: 99%

Glycolysis and Glutamate Accumulation into Synaptic Vesicles

Ikemoto

Bole

Ueda

2003

Journal of Biological Chemistry

174

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Glucose is the major source of brain energy and is essential for maintaining normal brain and neuronal function. Hypoglycemia causes impaired synaptic transmission. This occurs even before significant reduction in global cellular ATP concentration, and relationships among glycolysis, ATP supply, and synaptic transmission are not well understood. We demonstrate that the glycolytic enzymes glyceraldehyde phosphate dehydrogenase (GAPDH) and 3-phosphoglycerate kinase (3-PGK) are enriched in synaptic vesicles, forming a functional complex, and that synaptic vesicles are capable of accumulating the excitatory neurotransmitter glutamate by harnessing ATP produced by vesiclebound GAPDH/3-PGK at the expense of their substrates. The GAPDH inhibitor iodoacetate suppressed GAPDH/ 3-PGK-dependent, but not exogenous ATP-dependent, Glycolysis plays a vital role in maintaining normal brain function. Glucose is known to serve as the major substrate for cerebral energy under normal conditions (1). Recent evidence suggests a direct correlation between glucose utilization and cognitive function (2). Reduction of glucose levels results in pathophysiological states and abnormal electrophysiological activity; however, this occurs long before significant alteration in tissue ATP levels is detected (3-7). Substitution of pyruvate for glucose does not support normal evoked neuronal activity, although tissue ATP level returns to normal (8 -10). Abnormal synaptic transmission caused by hypoglycemia occurs in part if not entirely by a presynaptic mechanism (7,11,12). Fleck et al. (7) have shown that substantial reduction of extracellular glucose results in a decrease in stimulus-evoked Glu release, with no changes in ATP levels. These studies together suggest that glycolysis or glycolytic intermediate(s) are necessary for normal synaptic transmission independent of global cellular ATP levels.In an attempt to reveal the underlying mechanism of hypoglycemia-induced aberrant synaptic transmission, we previously explored the possibility that glycolytic intermediates could modify proteins localized in the nerve ending (13,14). 3-Phosphoglycerate (3-PG) 1 was demonstrated to stimulate phosphorylation of 155-and 72-kDa proteins. The latter was identified as glucose-1,6-bisphosphate synthetase, and 1,3-bisphosphoglycerate (1,3-BPG) was found to serve as the direct substrate for phosphorylation of this enzyme, by donating 1-phosphate. Both of these phosphorylated proteins are enriched in the synaptosomal (nerve ending preparation) as well as cell body soluble fractions, but the significance of these modifications in synaptic transmission remains unclear.In this paper, we show that the glycolytic intermediate 1,3-BPG forms an acyl-enzyme intermediate with vesicle-bound glyceraldehyde phosphate dehydrogenase (GAPDH), that vesicle-bound GAPDH exists in a complex with 3-phosphoglycerate kinase (3-PGK), and that activation of vesicle-associated GAPDH and 3-PGK is sufficient to support vesicular uptake of Glu. Glutamate is now recognized as the major ex...

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“…Assuming that cE, > CL 9 k -ilk1 (6) the ligand will be present almost exclusively as the EIL complex after preincubation with El. The concentration of free ligand will then remain low ([L] < cL) also after subsequent addition of E2, such that Eqn (3) reduces approximately to (1). which gives With the additional assumption that both enzymes are used in large excess to ligand, Eqns (4) and (5) …”

Section: Kinetic Measurementsmentioning

confidence: 99%

“…The kinetic evidence for direct metabolite transfer reported by Weber and Bernhard [1] was based mainly on observed effects of phosphoglycerate kinase on the catalytic interaction of glyceraldehyde-3-phosphate dehydrogenase with bisphosphoglycerate and NADH. Our main approach has been to examine the kinetics of the two-enzyme system in the presence of NAD' instead of NADH.…”

Section: Mechanism Of Bisphosphoglycerate Transfer In the Binding Expmentioning

confidence: 99%

Mechanism of 1,3‐bisphosphoglycerate transfer from phosphoglycerate kinase to glyceraldehyde‐3‐phosphate dehydrogenase

Kvassman

Pettersson

1989

European Journal of Biochemistry

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1. The kinetics of 1,3-bisphosphoglycerdte binding to glyceraldehyde-3-phosphate dehydrogenase have been examined by stopped-flow techniques in the absence and presence of phosphoglycerate kinase, using enzyme concentrations in the range 0.5 -40 pM. Kate and equilibrium constant estimates for the interaction of the ligand with the two enzymes are reported.2. The kinetics of ligand transfer from the binary complex of bisphosphoglycerate and phosphoglycerate kinase to the binary complex of NAD + and glyceraldehyde-3-phosphate dehydrogenase conform excellently to the predictions of a standard free-diffusion mechanism and exhibit no detectable contributions from a mechanism of direct (channelized) transfer of bisphosphoglycerate between the two enzymes.3. Previously reported evidence that the binary complex of bisphosphoglycerate and phosphoglycerate kinase may act (in the presence of NADH) as a substrate for glyceraldehyde-3-phosphate dehydrogenase according to Michaelis-Menten kinetics is based on a misinterpretation of the experimental observations that can be attributed to neglect of the autocatalytic effect of NAD + produced during the reaction. Experiments performed under conditions where the autocatalytic effect of NAD' is eliminated provide clear evidence that the kinetics of utilization of the kinase -bisphosphoglycerate complex for enzymic NADH reduction are consistent with prior dissociation of the complex according to a free-diffusion mechanism of metabolite transfer and incompatible with a mechanism of direct metabolite transfer.4. A kinetic argument is presented which renders implausible the very idea that direct metabolite transfer between 'soluble' consecutive enzymes in metabolic pathways may offer any catalytic advantages in comparison to metabolite transfer by free diffusion. A mechanism of direct metabolite transfer seems intuitively attractive only because one tends to disregard the diffusional processes required to bring the consecutive enzymes together and to separate them when the transfer has been completed. Direct metabolite transfer would be expected to be catalytically advantageous only in tightly bound multienzyme complexes showing no kinetically significant tendency to dissociate.5 . It is concluded that mechanisms of direct metabolite transfer have not been convincingly demonstrated to apply, nor are they likely to apply, between 'soluble' consecutive enzymes in metabolic pathways, at least not in the glycolytic sequence of reactions.Weber and Bernhard [l] in their kinetic study of the mechanism of 1,3-bisphosphoglycerate transfer from phosphoglycerate kinase to glyceraldehyde-3-phosphate dehydrogenase pointed out that concentrations of glycolytic enzymes in muscle cells usually exceed the concentrations of the substrates they act upon, such that many glycolytic intermediates may be present predominantly in an enzyme-bound form. The authors, therefore, designed a series of kinetic experiments to test if the tight binary complex formed between bisphosphoglycerate and phosphoglycerate ki...

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Transfer of 1,3-diphosphoglycerate between glyceraldehyde 3-phosphate dehydrogenase and 3-phosphoglycerate kinase via an enzyme-substrate-enzyme complex

Cited by 109 publications

References 22 publications

The Muscle-specific Calmodulin-dependent Protein Kinase Assembles with the Glycolytic Enzyme Complex at the Sarcoplasmic Reticulum and Modulates the Activity of Glyceraldehyde-3-phosphate Dehydrogenase in a Ca2+/Calmodulin-dependent Manner

The Muscle-specific Calmodulin-dependent Protein Kinase Assembles with the Glycolytic Enzyme Complex at the Sarcoplasmic Reticulum and Modulates the Activity of Glyceraldehyde-3-phosphate Dehydrogenase in a Ca2+/Calmodulin-dependent Manner

Glycolysis and Glutamate Accumulation into Synaptic Vesicles

Mechanism of 1,3‐bisphosphoglycerate transfer from phosphoglycerate kinase to glyceraldehyde‐3‐phosphate dehydrogenase

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