PDBsum entry: 1gy3

Transferase/transferase substrate

PDB id: 1gy3

Contents

Protein chains

296 a.a. *

258 a.a. *

Ligands

HIS-HIS-ALA-SER-PRO-ARG-LYS ×2

ATP ×2

NO3 ×2

GOL ×2

Metals

_MG ×2

Waters ×162

* Residue conservation analysis

PDB id:

1gy3

Name:

Transferase/transferase substrate

Title:

Pcdk2/cyclin a in complex with mgadp, nitrate and peptide su

Structure:

Cell division protein kinase 2. Chain: a, c. Synonym: p33 protein kinase. Engineered: yes. Other_details: phosphorylated on thr160. Cyclin a2. Chain: b, d. Fragment: residues 175-432. Engineered: yes.

Source:

Homo sapiens. Human. Organism_taxid: 9606. Expressed in: escherichia coli. Expression_system_taxid: 562. Other_details: cdk2 was co-expressed with cak1 - produce th phospho-cdk2. Synthetic: yes

Biol. unit:

Trimer (from PDB file)

Resolution:

2.70Å R-factor: 0.250 R-free: 0.313

Authors:

A.Cook,E.D.Lowe,E.D.Chrysina,V.T.Skamnaki,N.G.Oikonomakos,L.

Key ref:

A.Cook et al. (2002). Structural studies on phospho-CDK2/cyclin A bound to nitrate, a transition state analogue: implications for the protein kinase mechanism. Biochemistry, 41, 7301-7311. PubMed id: 12044161 DOI: 10.1021/bi0201724

Date:

19-Apr-02 Release date: 29-Apr-02

Protein chains

P24941  (CDK2_HUMAN) -  Cyclin-dependent kinase 2

Seq: 298 a.a.

Struc: 296 a.a.*

Protein chains

P20248  (CCNA2_HUMAN) -  Cyclin-A2

Seq: 432 a.a.

Struc: 258 a.a.

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: Chains A, C: E.C.2.7.11.22 - Cyclin-dependent kinase.
• Reaction: ATP + a protein = ADP + a phosphoprotein

ATP + protein = ADP (Bound ligand (Het Group name = ATP) corresponds exactly) + phosphoprotein

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

 Gene Ontology (GO) functional annotation 

• Cellular component: cyclin-dependent protein kinase holoenzyme complex (15 terms)
• Biological process: regulation of gene silencing (34 terms)
• Biochemical function: nucleotide binding (13 terms)

reference

DOI no: 10.1021/bi0201724

Biochemistry 41:7301-7311 (2002)

PubMed id: 12044161

Structural studies on phospho-CDK2/cyclin A bound to nitrate, a transition state analogue: implications for the protein kinase mechanism.

A.Cook, E.D.Lowe, E.D.Chrysina, V.T.Skamnaki, N.G.Oikonomakos, L.N.Johnson.

ABSTRACT

Eukaryotic protein kinases catalyze the phosphoryl transfer of the gamma-phosphate of ATP to the serine, threonine, or tyrosine residue of protein substrates. The catalytic mechanism of phospho-CDK2/cyclin A (pCDK2/cyclin A) has been probed with structural and kinetic studies using the trigonal NO(3)(-) ion, which can be viewed as a mimic of the metaphosphate transition state. The crystal structure of pCDK2/cyclin A in complex with Mg(2+)ADP, nitrate, and a heptapeptide substrate has been determined at 2.7 A. The nitrate ion is located between the beta-phosphate of ADP and the hydroxyl group of the serine residue of the substrate. In one molecule of the asymmetric unit, the nitrate is close to the beta-phosphate of ADP (distance from the nitrate nitrogen to the nearest beta-phosphate oxygen of 2.5 A), while in the other subunit, the nitrate is closer to the substrate serine (distance of 2.1 A). Kinetic studies demonstrate that nitrate is not an effective inhibitor of protein kinases, consistent with the structural results that show the nitrate ion makes few stabilizing interactions with CDK2 at the catalytic site. The binding of orthovanadate was also investigated as a mimic of a pentavalent phosphorane intermediate of an associative mechanism for phosphoryl transfer. No vanadate was observed bound in a 3.4 A resolution structure of pCDK2/cyclin A in the presence of Mg(2+)ADP, and vanadate did not inhibit the kinase reaction. The results support the notion that the protein kinase reaction proceeds through a mostly dissociative mechanism with a trigonal planar metaphosphate intermediate rather than an associative mechanism that involves a pentavalent phosphorane intermediate.