A phyloproteomic characterization of in vitro autophosphorylation in calcium-dependent protein kinases.
Calcium-dependent protein kinases (CDPKs) are a novel class of signaling molecules that have been broadly implicated in relaying specific calcium-mediated responses to biotic and abiotic stress as well as developmental cues in both plants and protists. Calcium-dependent autophosphorylation has been observed in almost all CDPKs examined, but a physiological role for autophosphorylation has not been demonstrated. To date, only a handful of autophosphorylation sites have been mapped to specific residues within CDPK amino acid sequences. In an attempt to gain further insight into this phenomenon, we have mapped autophosphorylation sites and compared these phosphorylation patterns among multiple CDPK isoforms. From eight CDPKs and two CDPK-related kinases from Arabidopsis thaliana and Plasmodium falciparum, 31 new autophosphorylation sites were characterized, which in addition to the previously described sites, allowed the identification of five conserved loci. Of the 35 total sites analyzed approximately one-half were observed in the N-terminal variable domain. Homology models were generated for the protein kinase and calmodulin-like domains, each containing two of the five conserved sites, to allow intelligent speculation regarding subsequent lines of investigation.
Hegeman
Adrian D
AD
University of Wisconsin Biotechnology Center, Madison, 53706, USA.
Rodriguez
Miguel
M
Han
Byung Woo
BW
Uno
Yuichi
Y
Phillips
George N
GN
Hrabak
Estelle M
EM
Cushman
John C
JC
Harper
Jeff F
JF
Harmon
Alice C
AC
Sussman
Michael R
MR
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry; D000818:Animals; D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics; D002118:Calcium / Q000378:metabolism; D017124:Conserved Sequence; D003627:Data Interpretation, Statistical; D004926:Escherichia coli / Q000235:genetics; D006868:Hydrolysis; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010766:Phosphorylation; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010944:Plants / Q000201:enzymology* / Q000235:genetics; D010963:Plasmodium falciparum / Q000201:enzymology / Q000235:genetics; D011494:Protein Kinases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D020543:Proteome / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D017871:Calcium-Calmodulin-Dependent Protein Kinases
D000596:Amino Acids; D002118:Calcium; D007527:Isoenzymes; D010940:Plant Proteins; D011494:Protein Kinases; D020543:Proteome; D017871:Calcium-Calmodulin-Dependent Protein Kinases