Phosphorylase kinase

 

Eukaryotic protein kinases catalyse the transfer of ATP-gamma phosphate to serine, threonine and tyrosine residues on specific target proteins. The family of protein kinases represents one of the largest protein superfamilies, and structures of active kinases show similar conformations in key regions involved in ATP and protein substrate recognition domains. Phosphorylase kinase, the first protein kinase to be discovered, catalyses the phosphorylation of a single serine residue, Ser14 of inactive phosphorylase B (GPb). Phosphorylation converts the protein to active glycogen phosphorylase A (GPa), which catalyses glycogen degredation.

 

Reference Protein and Structure

Sequence
P00518 UniProt (2.7.11.1, 2.7.11.19, 2.7.11.26) IPR002291 (Sequence Homologues) (PDB Homologues)
Biological species
Oryctolagus cuniculus (rabbit) Uniprot
PDB
2phk - THE CRYSTAL STRUCTURE OF A PHOSPHORYLASE KINASE PEPTIDE SUBSTRATE COMPLEX: KINASE SUBSTRATE RECOGNITION (2.6 Å) PDBe PDBsum 2phk
Catalytic CATH Domains
1.10.510.10 CATHdb (see all for 2phk)
Cofactors
Manganese(2+) (1), Magnesium(2+) (2) Metal MACiE
Click To Show Structure

Enzyme Reaction (EC:2.7.11.19)

L-serine residue
CHEBI:29999ChEBI
+
ATP(4-)
CHEBI:30616ChEBI
→
ADP(3-)
CHEBI:456216ChEBI
+
O-phospho-L-serine(2-) residue
CHEBI:83421ChEBI
+
hydron
CHEBI:15378ChEBI
Alternative enzyme names: Dephosphophosphorylase kinase, Glycogen phosphorylase kinase, Phosphorylase b kinase, Phosphorylase kinase (phosphorylating), Phosphorylase B kinase, STK17, PHK,

Enzyme Mechanism

Introduction

Asp149 deprotonates the protein substrate hydroxyl group, initiating nucleophilic attack upon the gamma-phosphate of ATP in a substitution reaction that eliminates ADP. The phosphorylated protein deprotonates Asp149 [PMID:10545198].

Catalytic Residues Roles

UniProt PDB* (2phk)
Lys152 Lys151(138)A Transition state stabiliser. hydrogen bond donor, electrostatic stabiliser
Thr187 Thr186(173)A Activates Asp149. activator, hydrogen bond donor
Asp150 Asp149(136)A Acts as a general acid/base catalyst. hydrogen bond acceptor, hydrogen bond donor, proton acceptor, proton donor
Asn155, Asp168 Asn154(141)A, Asp167(154)A Bind the Mg(II) ions. Asp168 binds both, Asp154 binds one only. metal ligand
*PDB label guide - RESx(y)B(C) - RES: Residue Name; x: Residue ID in PDB file; y: Residue ID in PDB sequence if different from PDB file; B: PDB Chain; C: Biological Assembly Chain if different from PDB. If label is "Not Found" it means this residue is not found in the reference PDB.

Chemical Components

proton transfer, bimolecular nucleophilic substitution, overall reactant used, overall product formed, intermediate formation, native state of enzyme regenerated, intermediate terminated

References

  1. Skamnaki VT et al. (1999), Biochemistry, 38, 14718-14730. Catalytic mechanism of phosphorylase kinase probed by mutational studies. DOI:10.2210/pdb1ql6/pdb. PMID:10545198.
  2. Lowe ED et al. (1997), EMBO J, 16, 6646-6658. The crystal structure of a phosphorylase kinase peptide substrate complex: kinase substrate recognition. DOI:10.1093/emboj/16.22.6646. PMID:9362479.
  3. Owen DJ et al. (1995), Structure, 3, 467-482. Two structures of the catalytic domain of phosphorylase kinase: an active protein kinase complexed with substrate analogue and product. DOI:10.1016/s0969-2126(01)00180-0. PMID:7663944.

Step 1. Asp149 deprotonates the protein substrate hydroxyl group, which initiates a nucleophilic attack upon the gamma-phosphate of ATP in a substitution reaction that eliminates ADP.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Asp149(136)A hydrogen bond acceptor
Lys151(138)A electrostatic stabiliser, hydrogen bond donor
Thr186(173)A hydrogen bond donor, activator
Asp167(154)A metal ligand
Asn154(141)A metal ligand
Asp149(136)A proton acceptor

Chemical Components

proton transfer, ingold: bimolecular nucleophilic substitution, overall reactant used, overall product formed, intermediate formation

Step 2. The phosphorylated protein deprotonates Asp149.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Asp149(136)A hydrogen bond donor, hydrogen bond acceptor
Lys151(138)A electrostatic stabiliser, hydrogen bond donor
Thr186(173)A hydrogen bond donor
Asp167(154)A metal ligand
Asn154(141)A metal ligand
Asp149(136)A proton donor

Chemical Components

overall product formed, native state of enzyme regenerated, intermediate terminated, proton transfer
Download: Image, Marvin File

Step 1. Asp149 deprotonates the protein substrate hydroxyl group, which initiates a nucleophilic attack upon the gamma-phosphate of ATP in a substitution reaction that eliminates ADP.

Step 2. The phosphorylated protein deprotonates Asp149.

Products.

Contributors

Gemma L. Holliday, Gail J. Bartlett, Daniel E. Almonacid, Sophie T. Williams, Craig Porter