Strength in numbers: Phosphofructokinase polymerization prevails in the liver

Zaganjor, Elma; Spinelli, Jessica B.; Haigis, Marcia C.

doi:10.1083/jcb.201706005

Cited by 4 publications

(1 citation statement)

References 14 publications

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“…Only recently has filament formation by non-ATP or GTPases been appreciated. New imaging technologies have allowed for large-scale screening of protein localization in cells and have revealed filament formation by many metabolic and other enzymes, sometimes coinciding with particular phases of the cell cycle, with certain stress conditions, or as part of signaling pathways (1)(2)(3)(4)(5)(6)(7)(22)(23)(24)(25)(26)(27)(28)(29). Run-on oligomers, or ROO filaments, are by definition filaments, although not all ROO filaments form filaments as large and as stable as others.…”

Section: Discussionmentioning

confidence: 99%

The run-on oligomer filament enzyme mechanism of SgrAI: Part 2. Kinetic modeling of the full DNA cleavage pathway

Park

Sanchez

Barahona

et al. 2018

Journal of Biological Chemistry

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Filament or run-on oligomer formation by enzymes is now recognized as a widespread phenomenon with potentially unique enzyme regulatory properties and biological roles. SgrAI is an allosteric type II restriction endonuclease that forms run-on oligomeric filaments with activated DNA cleavage activity and altered DNA sequence specificity. In this two-part work, we measure individual steps in the run-on oligomer filament mechanism to address specific questions of cooperativity, trapping, filament growth mechanisms, and sequestration of activity using fluorophore-labeled DNA, kinetic FRET measurements, and reaction modeling with global data fitting. The final models and rate constants show that the assembly step involving association of SgrAI-DNA complexes into the run-on oligomer filament is relatively slow (3-4 orders of magnitude slower than diffusion limited) and rate-limiting at low to moderate concentrations of SgrAI-DNA. The disassembly step involving dissociation of complexes of SgrAI-DNA from each other in the run-on oligomer filament is the next slowest step but is fast enough to limit the residence time of any one copy of SgrAI or DNA within the dynamic filament. Further, the rate constant for DNA cleavage is found to be 4 orders of magnitude faster in the run-on oligomer filament than in isolated SgrAI-DNA complexes and faster than dissociation of SgrAI-DNA complexes from the run-on oligomer filament, making the reaction efficient in that each association into the filament likely leads to DNA cleavage before filament dissociation.

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Section: Discussionmentioning

confidence: 99%

The run-on oligomer filament enzyme mechanism of SgrAI: Part 2. Kinetic modeling of the full DNA cleavage pathway

Park

Sanchez

Barahona

et al. 2018

Journal of Biological Chemistry

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Winter is coming: Regulation of cellular metabolism by enzyme polymerization in dormancy and disease

Montrose

Cabezas

Paukštytė

et al. 2020

Experimental Cell Research

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The run-on oligomer filament enzyme mechanism of SgrAI: Part 1. Assembly kinetics of the run-on oligomer filament

Park

Sanchez

Barahona

et al. 2018

Journal of Biological Chemistry

View full text Add to dashboard Cite

Filament or run-on oligomer formation by metabolic enzymes is now recognized as a widespread phenomenon having potentially unique enzyme regulatory properties and biological roles, and its dysfunction is implicated in human diseases such as cancer, diabetes, and developmental disorders. SgrAI is a bacterial allosteric type II restriction endonuclease that binds to invading phage DNA, may protect the host DNA from off-target cleavage activity, and forms run-on oligomeric filaments with enhanced DNA-cleavage activity and altered DNA sequence specificity. However, the mechanisms of SgrAI filament growth, cooperativity in filament formation, sequestration of enzyme activity, and advantages over other filament mechanisms remain unknown. In this first of a two-part series, we developed methods and models to derive association and dissociation rate constants of DNA-bound SgrAI in run-on oligomers and addressed the specific questions of cooperativity and filament growth mechanisms. We show that the derived rate constants are consistent with the run-on oligomer sizes determined by EM analysis and are most consistent with a noncooperative growth mode of the run-on oligomer. These models and methods are extended in the accompanying article to include the full DNA-cleavage pathway and address specific questions related to the run-on oligomer mechanism including the sequestration of DNA-cleavage activity and trapping of products.

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Strength in numbers: Phosphofructokinase polymerization prevails in the liver

Abstract: Zaganjor et al. preview work from Webb et al. that shows that the liver-specific isoform of the glycolysis enzyme PFK1 can assemble into filaments and localize to distinct puncta near the plasma membrane.

Cited by 4 publications

References 14 publications

The run-on oligomer filament enzyme mechanism of SgrAI: Part 2. Kinetic modeling of the full DNA cleavage pathway

The run-on oligomer filament enzyme mechanism of SgrAI: Part 2. Kinetic modeling of the full DNA cleavage pathway

Winter is coming: Regulation of cellular metabolism by enzyme polymerization in dormancy and disease

The run-on oligomer filament enzyme mechanism of SgrAI: Part 1. Assembly kinetics of the run-on oligomer filament

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