PDBsum entry 6wp5

Transferase

PDB id: 6wp5

Contents

Protein chains

510 a.a.

407 a.a.

Ligands

GOL ×9

OXL ×4

FBP ×3

Metals

__K ×4

_MG ×3

Waters ×209

PDB id:

6wp5

Name:

Transferase

Title:

Pyruvate kinase m2 mutant-s37d

Structure:

Pyruvate kinase pkm. Chain: b, a, c, d. Synonym: cytosolic thyroid hormone-binding protein,cthbp,opa- interacting protein 3,oip-3,pyruvate kinase 2/3,pyruvate kinase muscle isozyme,thyroid hormone-binding protein 1,thbp1,tumor m2-pk, p58. Engineered: yes. Mutation: yes

Source:

Homo sapiens. Human. Organism_taxid: 9606. Gene: pkm, oip3, pk2, pk3, pkm2. Expressed in: escherichia coli. Expression_system_taxid: 562

Resolution:

2.17Å R-factor: 0.219 R-free: 0.260

Authors:

S.Nandi,M.Razzaghi,D.Srivastava,M.Dey

Key ref:

S.Nandi et al. (2020). Structural basis for allosteric regulation of pyruvate kinase M2 by phosphorylation and acetylation. J Biol Chem, 295, 17425-17440. PubMed id: 33453989 DOI: 10.1074/jbc.RA120.015800

Date:

26-Apr-20 Release date: 30-Sep-20

Protein chains

P14618  (KPYM_HUMAN) -  Pyruvate kinase PKM from Homo sapiens

Seq: 531 a.a.

Struc: 510 a.a.*

Protein chain

P14618  (KPYM_HUMAN) -  Pyruvate kinase PKM from Homo sapiens

Seq: 531 a.a.

Struc: 407 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

Enzyme reactions

• Enzyme class: Chains B, A, C, D: E.C.2.7.1.40 - pyruvate kinase.
• Reaction: pyruvate + ATP = phosphoenolpyruvate + ADP + H⁺

pyruvate + ATP (Bound ligand (Het Group name = GOL) matches with 71.43% similarity) = phosphoenolpyruvate + ADP + H(+)

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1074/jbc.RA120.015800

J Biol Chem 295:17425-17440 (2020)

PubMed id: 33453989

Structural basis for allosteric regulation of pyruvate kinase M2 by phosphorylation and acetylation.

S.Nandi, M.Razzaghi, D.Srivastava, M.Dey.

ABSTRACT

Pyruvate kinase muscle isoform 2 (PKM2) is a key glycolytic enzyme and transcriptional coactivator and is critical for tumor metabolism. In cancer cells, native tetrameric PKM2 is phosphorylated or acetylated, which initiates a switch to a dimeric/monomeric form that translocates into the nucleus, causing oncogene transcription. However, it is not known how these post-translational modifications (PTMs) disrupt the oligomeric state of PKM2. We explored this question via crystallographic and biophysical analyses of PKM2 mutants containing residues that mimic phosphorylation and acetylation. We find that the PTMs elicit major structural reorganization of the fructose 1,6-bisphosphate (FBP), an allosteric activator, binding site, impacting the interaction with FBP and causing a disruption in oligomerization. To gain insight into how these modifications might cause unique outcomes in cancer cells, we examined the impact of increasing the intracellular pH (pH_i) from ∼7.1 (in normal cells) to ∼7.5 (in cancer cells). Biochemical studies of WT PKM2 (wtPKM2) and the two mimetic variants demonstrated that the activity decreases as the pH is increased from 7.0 to 8.0, and wtPKM2 is optimally active and amenable to FBP-mediated allosteric regulation at pH_i 7.5. However, the PTM mimetics exist as a mixture of tetramer and dimer, indicating that physiologically dimeric fraction is important and might be necessary for the modified PKM2 to translocate into the nucleus. Thus, our findings provide insight into how PTMs and pH regulate PKM2 and offer a broader understanding of its intricate allosteric regulation mechanism by phosphorylation or acetylation.