PDBsum entry 2phk

Complex (transferase/peptide)

PDB id: 2phk

Contents

Protein chain

277 a.a. *

Ligands

ARG-GLN-MET-SER-PHE-ARG-LEU

ATP

GOL

Metals

_MN ×2

Waters ×86

* Residue conservation analysis

PDB id:

2phk

Name:

Complex (transferase/peptide)

Title:

The crystal structure of a phosphorylase kinase peptide substrate complex: kinase substrate recognition

Structure:

Phosphorylase kinase. Chain: a. Fragment: catalytic domain. Engineered: yes. Mc-peptide. Chain: b. Engineered: yes

Source:

Oryctolagus cuniculus. Rabbit. Organism_taxid: 9986. Strain: new zealand white. Cell_line: bl21. Organ: skeletal. Tissue: skeletal muscle. Gene: phkg. Expressed in: escherichia coli bl21(de3).

Biol. unit:

Tetramer (from PQS)

Resolution:

2.60Å R-factor: 0.253 R-free: 0.300

Authors:

E.D.Lowe,M.E.M.Noble,V.T.Skamnaki,N.G.Oikonomakos,D.J.Owen, L.N.Johnson

Key ref:

E.D.Lowe et al. (1997). The crystal structure of a phosphorylase kinase peptide substrate complex: kinase substrate recognition. EMBO J, 16, 6646-6658. PubMed id: 9362479 DOI: 10.1093/emboj/16.22.6646

Date:

18-Jun-98 Release date: 13-Jan-99

Protein chain

P02853  (PHSB_PHAVU) -  Phaseolin, beta-type from Phaseolus vulgaris

Seq: 421 a.a.

Struc: 277 a.a.*

* PDB and UniProt seqs differ at 242 residue positions (black crosses)

Enzyme reactions

• Enzyme class 1: E.C.2.7.11.1 - non-specific serine/threonine protein kinase.
• Reaction:
  1. L-seryl-[protein] + ATP = O-phospho-L-seryl-[protein] + ADP + H⁺
  2. L-threonyl-[protein] + ATP = O-phospho-L-threonyl-[protein] + ADP + H⁺

L-seryl-[protein] (Bound ligand (Het Group name = ATP) corresponds exactly) + ATP = O-phospho-L-seryl-[protein] + ADP + H(+)

L-threonyl-[protein] (Bound ligand (Het Group name = ATP) corresponds exactly) + ATP = O-phospho-L-threonyl-[protein] + ADP + H(+)

• Enzyme class 2: E.C.2.7.11.19 - phosphorylase kinase.
• Reaction: 2 ATP + phosphorylase b = 2 ADP + phosphorylase a

2 × ATP (Bound ligand (Het Group name = ATP) corresponds exactly) + phosphorylase b = 2 × ADP + phosphorylase a

• Cofactor: Ca(2+)
• Enzyme class 3: E.C.2.7.11.26 - [tau protein] kinase.
• Reaction:
  1. L-seryl-[tau protein] + ATP = O-phospho-L-seryl-[tau protein] + ADP + H⁺
  2. L-threonyl-[tau protein] + ATP = O-phospho-L-threonyl-[tau protein] + ADP + H⁺

2 × L-seryl-[tau protein] (Bound ligand (Het Group name = ATP) corresponds exactly) + ATP = 2 × O-phospho-L-seryl-[tau protein] + ADP + H(+)

L-threonyl-[tau protein] (Bound ligand (Het Group name = ATP) corresponds exactly) + ATP = O-phospho-L-threonyl-[tau protein] + ADP + H(+)

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

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

reference

DOI no: 10.1093/emboj/16.22.6646

EMBO J 16:6646-6658 (1997)

PubMed id: 9362479

The crystal structure of a phosphorylase kinase peptide substrate complex: kinase substrate recognition.

E.D.Lowe, M.E.Noble, V.T.Skamnaki, N.G.Oikonomakos, D.J.Owen, L.N.Johnson.

ABSTRACT

The structure of a truncated form of the gamma-subunit of phosphorylase kinase (PHKgammat) has been solved in a ternary complex with a non-hydrolysable ATP analogue (adenylyl imidodiphosphate, AMPPNP) and a heptapeptide substrate related in sequence to both the natural substrate and to the optimal peptide substrate. Kinetic characterization of the phosphotransfer reaction confirms the peptide to be a good substrate, and the structure allows identification of key features responsible for its high affinity. Unexpectedly, the substrate peptide forms a short anti-parallel beta-sheet with the kinase activation segment, the region which in other kinases plays an important role in regulation of enzyme activity. This anchoring of the main chain of the substrate peptide at a fixed distance from the gamma-phosphate of ATP explains the selectivity of PHK for serine/threonine over tyrosine as a substrate. The catalytic core of PHK exists as a dimer in crystals of the ternary complex, and the relevance of this phenomenon to its in vivo recognition of dimeric glycogen phosphorylase b is considered.

Selected figure(s)

Figure 3.
Figure 3 Conformations of kinase inhibitor and substrate peptides. (A) Conformation of residues 11 -17 of GPa. (B) Conformation of residues 11 -17 of GPb. (C) Conformation of the MC-peptide, as observed in complex with PHK [t]. (D) Conformation of the equivalent part of the protein kinase inhibitor peptide, as observed in complex with cAPK.

Figure 6.
Figure 6 Interactions of nucleophile, base and phosphate group. Interactions which might occur in a general base-catalysed reaction mechanism via: (A) the observed structure of the ternary complex and (B) an alternative conformation modelled by giving the attacking serine residue a 1 angle of -60°.

The above figures are reprinted from an Open Access publication published by Macmillan Publishers Ltd: EMBO J (1997, 16, 6646-6658) copyright 1997.

Figures were selected by an automated process.