Pim kinase substrate identification and specificity.
The Pim family of Ser/Thr kinases has been implicated in the process of lymphomagenesis and cell survival. Known substrates of Pim kinases are few and poorly characterized. In this study we set out to identify novel Pim-2 substrates using the Kinase Substrate Tracking and Elucidation (KESTREL) approach. Two potential substrates, eukaryotic initiation factor 4B (eIF4B) and apoptosis inhibitor 5 (API-5), were identified from rat thymus extracts. Sequence comparison of the Pim-2 kinase phosphorylation sites of eIF4B and mouse BAD, the only other known Pim-2 substrate, revealed conserved amino acids preceding the phosphorylated serine residue. Stepwise replacement of the conserved residues produced a consensus sequence for Pim kinase recognition: RXRHXS. Pim-1 and Pim-2 catalyzed the phosphorylation of this recognition sequence 20-fold more efficiently than the original (K/R-K/R-R-K/R-L-S/T-a; a = small chain amino acid) Pim-1 phosphorylation site. The identification of the novel Pim kinase consensus sequence provides a more sensitive and versatile peptide based assay for screening modulators of Pim kinase activity.
Peng
Charline
C
Boehringer Ingelheim Pharmaceuticals, Inc. 900 Ridgebury Road, Ridgefield, CT 06877, USA.
Knebel
Axel
A
Morrice
Nick A
NA
Li
Xiang
X
Barringer
Kevin
K
Li
Jun
J
Jakes
Scott
S
Werneburg
Brian
B
Wang
Lian
L
D000595:Amino Acid Sequence; D000818:Animals; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D016384:Consensus Sequence; D039642:Eukaryotic Initiation Factors / Q000378:metabolism; D006801:Humans; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051573:Proto-Oncogene Proteins c-pim-1 / Q000378:metabolism*; D051381:Rats; D013379:Substrate Specificity; D013950:Thymus Gland / Q000378:metabolism
D051017:Apoptosis Regulatory Proteins; D039642:Eukaryotic Initiation Factors; C490587:PIM2 protein, human; D011518:Proto-Oncogene Proteins; C031799:eIF-4B; C496634:PIM1 protein, human; D017346:Protein Serine-Threonine Kinases; D051573:Proto-Oncogene Proteins c-pim-1