Structure function analysis of ADP-dependent cyanobacterial phosphofructokinase reveals new phylogenetic grouping in the PFK-A family

Lu Shen; Carmen Peraglie; David Podlesainski; Christina Stracke; Ravi Shankar Ojha; Frauke Caliebe; Markus Kaiser; Karl Forchhammer; Martin Hagemann; Kirstin Gutekunst; Jacky L Snoep; Christopher Bräsen; Bettina Siebers

doi:10.1016/j.jbc.2024.107868

Structure function analysis of ADP-dependent cyanobacterial phosphofructokinase reveals new phylogenetic grouping in the PFK-A family

J Biol Chem. 2024 Oct 9:107868. doi: 10.1016/j.jbc.2024.107868. Online ahead of print.

Affiliations

¹ Molecular Enzyme Technology and Biochemistry (MEB), Environmental Microbiology and Biotechnology (EMB), University of Duisburg-Essen, Universitätsstr. 5, Essen, 45141, State, Germany.
² Chemical Biology, Centre of Medical Biotechnology (ZMB), Faculty of Biology, University of Duisburg-Essen, Universitätsstr. 2, 45117 Essen, Germany.
³ Molekulare Pflanzenphysiologie, University of Kassel, Heinrich-Plett-Strasse 40, 34132 Kassel, Germany.
⁴ Microbiology, University of Tübingen, Auf der Morgenstelle 28, 72076 Tübingen, Germany.
⁵ Plant Physiology, University of Rostock, Albert-Einstein-Str. 3, 18059 Rostock, Germany.
⁶ Biochemistry, University of Stellenbosch, Private Bag X1, 7600 Stellenbosch, South Africa; Molecular Cell Biology, Vrije Universiteit Amsterdam, De Boelelaan 1108, 1081 HZ Amsterdam, The Netherlands.
⁷ Molecular Enzyme Technology and Biochemistry (MEB), Environmental Microbiology and Biotechnology (EMB), University of Duisburg-Essen, Universitätsstr. 5, Essen, 45141, State, Germany. Electronic address: [email protected].
⁸ Molecular Enzyme Technology and Biochemistry (MEB), Environmental Microbiology and Biotechnology (EMB), University of Duisburg-Essen, Universitätsstr. 5, Essen, 45141, State, Germany. Electronic address: [email protected].

PMID: 39393572
DOI: 10.1016/j.jbc.2024.107868

Abstract

Dependent on the light conditions, photosynthetic organisms switch between carbohydrate synthesis or breakdown, for which the reversibility of carbohydrate metabolism, including glycolysis, is essential. Kinetic regulation of phosphofructokinase (PFK), a key-control point in glycolysis, was studied in the cyanobacterium Synechocystis sp. PCC 6803. The two PFK iso-enzymes (PFK- A1, PFK-A2), were found to use ADP instead of ATP, and have similar kinetic characteristics, but differ in their allosteric regulation. PFK-A1 is inhibited by 3- phosphoglycerate, a product of the Calvin-Benson-Bassham cycle, while PFK-A2 is inhibited by ATP, which is provided by photosynthesis. This regulation enables cyanobacteria to switch PFK off in light, and on in darkness. ADP dependence has not been reported before for the PFK-A enzyme family, and was thought to be restricted to the PFK-B ribokinase superfamily. Phosphate donor specificity within the PFK-A family could be related to specific binding motifs in ATP-, ADP-, and PPi-dependent PFK-As. Phylogenetic analysis revealed a distribution of ADP-PFK-As in cyanobacteria and in a few alphaproteobacteria, which was confirmed in enzymatic studies.

Keywords: ADP; ADP-dependent PFK-A; ATP; Cyanobacteria; PFK-A superfamily; PP(i) signature binding motifs; allosteric regulation.