title: Insights into cyclin groove recognition: complex crystal structures and inhibitor design through ligand exchange.
abstract: Inhibition of CDK2/CA (cyclin-dependent kinase 2/cyclin A complex) activity through blocking of the substrate recognition site in the cyclin A subunit has been demonstrated to be an effective method for inducing apoptosis in tumor cells. We have used the cyclin binding motif (CBM) present in the tumor suppressor proteins p21(WAF1) and p27(KIP1) as a template to optimize the minimal sequence necessary for CDK2/CA inhibition. A series of peptides were prepared, containing nonnatural amino acids, which possess nano- to micromolar CDK2-inhibitory activity. Here we present X-ray structures of the protein complex CDK2/CA, together with the cyclin groove-bound peptides H-Ala-Ala-Abu-Arg-Ser-Leu-Ile-(p-F-Phe)-NH(2) (peptide 1), H-Arg-Arg-Leu-Ile-Phe-NH(2) (peptide 2), Ac-Arg-Arg-Leu-Asn-(m-Cl-Phe)-NH(2) (peptide 3), H-Arg-Arg-Leu-Asn-(p-F-Phe)-NH(2) (peptide 4), and H-Cit-Cit-Leu-Ile-(p-F-Phe)-NH(2) (peptide 5). Some of the peptide complexes presented here were obtained through the novel technique of ligand exchange within protein crystals. This method may find general application for obtaining complex structures of proteins with surface-bound ligands.
authors.affiliation: Cyclacel Ltd., James Lindsay Place, Dundee DD1 5JJ, Scotland, United Kingdom. gkontopidis@cyclacel.com
authors.forename: George
authors.initials: G
authors.lastname: Kontopidis
authors.forename: Martin J I
authors.initials: MJ
authors.lastname: Andrews
authors.forename: Campbell
authors.initials: C
authors.lastname: McInnes
authors.forename: Angela
authors.initials: A
authors.lastname: Cowan
authors.forename: Helen
authors.initials: H
authors.lastname: Powers
authors.forename: Lorraine
authors.initials: L
authors.lastname: Innes
authors.forename: Andy
authors.initials: A
authors.lastname: Plater
authors.forename: Gary
authors.initials: G
authors.lastname: Griffiths
authors.forename: Dougie
authors.initials: D
authors.lastname: Paterson
authors.forename: Daniella I
authors.initials: DI
authors.lastname: Zheleva
authors.forename: David P
authors.initials: DP
authors.lastname: Lane
authors.forename: Stephen
authors.initials: S
authors.lastname: Green
authors.forename: Malcolm D
authors.initials: MD
authors.lastname: Walkinshaw
authors.forename: Peter M
authors.initials: PM
authors.lastname: Fischer
mesh_terms: D020816:Amino Acid Motifs; D001665:Binding Sites; D042846:CDC2-CDC28 Kinases / Q000037:antagonists & inhibitors / Q000737:chemistry; D018360:Crystallography, X-Ray; D019925:Cyclin A / Q000037:antagonists & inhibitors / Q000737:chemistry; D051357:Cyclin-Dependent Kinase 2; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000737:chemistry*; D006801:Humans; D008024:Ligands; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D013329:Structure-Activity Relationship
chemical_list: C493956:CDKN1A protein, human; D019925:Cyclin A; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D008024:Ligands; D010455:Peptides; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2