8STX

Crystal structure of the F337A mutation of Trypanosoma cruzi glucokinase in the apo form (open conformation)

Summary for 8STX

• Entry DOI: 10.2210/pdb8stx/pdb
• Descriptor: Glucokinase (2 entities in total)
• Functional Keywords: glucokinase, transferase, mutation
• Biological source: Trypanosoma cruzi
• Total number of polymer chains: 2
• Total formula weight: 84301.11

Authors

Abiskaroon, B.,Carey, S.M.,D'Antonio, E.L.,Chruszcz, M. (deposition date: 2023-05-11, release date: 2023-05-31, Last modification date: 2023-12-13)

Primary citation

Carey, S.M.,Kearns, S.P.,Millington, M.E.,Buechner, G.S.,Alvarez Jr., B.E.,Daneshian, L.,Abiskaroon, B.,Chruszcz, M.,D'Antonio, E.L.
At the outer part of the active site in Trypanosoma cruzi glucokinase: The role of phenylalanine 337.
Biochimie, 218:8-19, 2023

PubMed Abstract: The hole mutagenesis approach was used to interrogate the importance of F337 in Trypanosoma cruzi glucokinase (TcGlcK) in order to understand the complete set of binding interactions that are made by d-glucosamine analogue inhibitors containing aromatic tail groups that can extend to the outer part of the active site. An interesting inhibitor of this analogue class includes 2-N-carboxybenzyl-2-deoxy-d-glucosamine (CBZ-GlcN), which exhibits strong TcGlcK binding with a K of 710 nM. The residue F337 is found at the outer part of the active site that stems from the second protein subunit of the homodimeric assembly. In this study, F337 was changed to leucine and alanine so as to diminish phenylalanine's side chain size and attenuate intermolecular interactions in this region of the binding cavity. Results from enzyme - inhibitor assays revealed that the phenyl group of F337 made dominant hydrophobic interactions with the phenyl group of CBZ-GlcN as opposed to π - π stacking interactions. Moreover, enzymatic activity assays and X-ray crystallographic experiments indicated that each of these site-directed mutants primarily retained their activity and had high structural similarity of their protein fold. A computed structure model of T. cruzi hexokinase (TcHxK), which was produced by the artificial intelligence system AlphaFold, was compared to an X-ray crystal structure of TcGlcK. Our structural analysis revealed that TcHxK lacked an F337 counterpart residue and probably exists in the monomeric form. We proposed that the d-glucosamine analogue inhibitors that are structurally similar to CBZ-GlcN may not bind as strongly in TcHxK as they do in TcGlcK because of absent van der Waals contact from residue side chains.

PubMed: 37741546
DOI: 10.1016/j.biochi.2023.09.014

Experimental method

X-RAY DIFFRACTION (2.4 Å)