Loss of stability and unfolding cooperativity in hPGK1 upon gradual structural perturbation of its N-terminal domain hydrophobic core

0564237 - MBÚ 2023 RIV DE eng J - Journal Article
Luis Pacheco-Garcia, J. - Loginov, Dmitry Sergej - Naganathan, A. N. - Vaňková, Pavla - Cano-Munoz, M. - Man, Petr - Pey, Angel L.
Loss of stability and unfolding cooperativity in hPGK1 upon gradual structural perturbation of its N-terminal domain hydrophobic core.
Scientific Reports. Roč. 12, č. 1 (2022), č. článku 17200. ISSN 2045-2322. E-ISSN 2045-2322
R&D Projects: GA MŠMT(CZ) ED1.1.00/02.0109
EU Projects: European Commission(XE) 731077 - EU_FT-ICR_MS
Research Infrastructure: CIISB II - 90127
Institutional support: RVO:61388971 ; RVO:86652036
Keywords : muscle 3-phosphoglycerate kinase * phosphoglycerate kinase * phosphoglycerate kinase * protein stability * kinetic stability * molecular-basis * denaturation * conformation * flexibility * modulation * ensembles
OECD category: Biochemistry and molecular biology
Impact factor: 4.6, year: 2022
Method of publishing: Open access
https://www.nature.com/articles/s41598-022-22088-1

Phosphoglycerate kinase has been a model for the stability, folding cooperativity and catalysis of a two-domain protein. The human isoform 1 (hPGK1) is associated with cancer development and rare genetic diseases that affect several of its features. To investigate how mutations affect hPGK1 folding landscape and interaction networks, we have introduced mutations at a buried site in the N-terminal domain (F25 mutants) that either created cavities (F25L, F25V, F25A), enhanced conformational entropy (F25G) or introduced structural strain (F25W) and evaluated their effects using biophysical experimental and theoretical methods. All F25 mutants folded well, but showed reduced unfolding cooperativity, kinetic stability and altered activation energetics according to the results from thermal and chemical denaturation analyses. These alterations correlated well with the structural perturbation caused by mutations in the N-terminal domain and the destabilization caused in the interdomain interface as revealed by H/D exchange under native conditions. Importantly, experimental and theoretical analyses showed that these effects are significant even when the perturbation is mild and local. Our approach will be useful to establish the molecular basis of hPGK1 genotype-phenotype correlations due to phosphorylation events and single amino acid substitutions associated with disease.
Permanent Link: https://hdl.handle.net/11104/0335944