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Kinetic and structural studies of Trypanosoma and Leishmania phosphofructokinases show evolutionary divergence and identify AMP as a switch regulating glycolysis versus gluconeogenesis

DOI: 10.1111/febs.15177 DOI Help

Authors: Peter M. Fernandes (The University of Edinburgh) , James Kinkead (The University of Edinburgh) , Iain W. Mcnae (The University of Edinburgh) , Monserrat Vasquez-Valdivieso (The University of Edinburgh) , Martin A. Wear (The University of Edinburgh) , Paul A. M. Michels (The University of Edinburgh) , Malcolm D. Walkinshaw (The University of Edinburgh)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: The Febs Journal , VOL 160

State: Published (Approved)
Published: January 2020
Diamond Proposal Number(s): 1225

Open Access Open Access

Abstract: Trypanosomatids possess glycosome organelles that contain much of the glycolytic machinery, including phosphofructokinase (PFK). We present kinetic and structural data for PFK from three human pathogenic trypanosomatids, illustrating intriguing differences that may reflect evolutionary adaptations to differing ecological niches. The activity of Leishmania PFK – to a much larger extent than Trypanosoma PFK – is reliant on AMP for activity regulation, with 1 mm AMP increasing the L. infantum PFK (LiPFK) kcat/K0.5F6P value by 10‐fold, compared to only a 1.3‐ and 1.4‐fold increase for T. cruzi and T. brucei PFK, respectively. We also show that Leishmania PFK melts at a significantly lower (> 15 °C) temperature than Trypanosoma PFKs and that addition of either AMP or ATP results in a marked stabilization of the protein. Sequence comparisons of Trypanosoma spp. and Leishmania spp. show that divergence of the two genera involved amino acid substitutions that occur in the enzyme’s ‘reaching arms’ and ‘embracing arms’ that determine tetramer stability. The dramatic effects of AMP on Leishmania activity compared with the Trypanosoma PFKs may be explained by differences between the T‐to‐R equilibria for the two families, with the low‐melting Leishmania PFK favouring the flexible inactive T‐state in the absence of AMP. Sequence comparisons along with the enzymatic and structural data presented here also suggest there was a loss of AMP‐dependent regulation in Trypanosoma species rather than gain of this characteristic in Leishmania species and that AMP acts as a key regulator in Leishmania governing the balance between glycolysis and gluconeogenesis.

Journal Keywords: allostery; glycolysis; Leishmania; phosphofructokinase; Trypanosoma

Diamond Keywords: Sleeping Sickness; Leishmaniasis; Enzymes

Subject Areas: Biology and Bio-materials, Chemistry

Instruments: I04-1-Macromolecular Crystallography (fixed wavelength)

Added On: 08/01/2020 14:04


Discipline Tags:

Pathogens Infectious Diseases Disease in the Developing World Health & Wellbeing Biochemistry Chemistry Structural biology Life Sciences & Biotech Parasitology

Technical Tags:

Diffraction Macromolecular Crystallography (MX)