title: Utilization of oriented peptide libraries to identify substrate motifs selected by ATM.
abstract: The ataxia telangiectasia mutated (ATM) gene encodes a serine/threonine protein kinase that plays a critical role in genomic surveillance and development. Here, we use a peptide library approach to define the in vitro substrate specificity of ATM kinase activity. The peptide library analysis identified an optimal sequence with a central core motif of LSQE that is preferentially phosphorylated by ATM. The contributions of the amino acids surrounding serine in the LSQE motif were assessed by utilizing specific peptide libraries or individual peptide substrates. All amino acids comprising the LSQE sequence were critical for maximum peptide substrate suitability for ATM. The DNA-dependent protein kinase (DNA-PK), a Ser/Thr kinase related to ATM and important in DNA repair, was compared with ATM in terms of peptide substrate selectivity. DNA-PK was found to be unique in its preference of neighboring amino acids to the phosphorylated serine. Peptide library analyses defined a preferred amino acid motif for ATM that permits clear distinctions between ATM and DNA-PK kinase activity. Data base searches using the library-derived ATM sequence identified previously characterized substrates of ATM, as well as novel candidate substrate targets that may function downstream in ATM-directed signaling pathways.
authors.affiliation: Center for Blood Research, Department of Pediatrics, Children's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.
authors.forename: T
authors.initials: T
authors.lastname: O'Neill
authors.forename: A J
authors.initials: AJ
authors.lastname: Dwyer
authors.forename: Y
authors.initials: Y
authors.lastname: Ziv
authors.forename: D W
authors.initials: DW
authors.lastname: Chan
authors.forename: S P
authors.initials: SP
authors.lastname: Lees-Miller
authors.forename: R H
authors.initials: RH
authors.lastname: Abraham
authors.forename: J H
authors.initials: JH
authors.lastname: Lai
authors.forename: D
authors.initials: D
authors.lastname: Hill
authors.forename: Y
authors.initials: Y
authors.lastname: Shiloh
authors.forename: L C
authors.initials: LC
authors.lastname: Cantley
authors.forename: G A
authors.initials: GA
authors.lastname: Rathbun
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D008969:Molecular Sequence Data; D019151:Peptide Library; D019869:Phosphatidylinositol 3-Kinases / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism*; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured; D025521:Tumor Suppressor Proteins
chemical_list: D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D019151:Peptide Library; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D025521:Tumor Suppressor Proteins