Purification and characterization of wheat and pine small basic protein substrates for plant calcium-dependent protein kinase.
A wheat basic protein (WBP) was purified to homogeneity from wheat germ by a protocol involving extraction, centrifugation, batchwise elution from carboxymethylcellulose (CM-52), acidification with trifluoroacetic acid, neutralization and HPLC on a SP5PW cation exchange column. WBP is a 10 kDa protein and is phosphorylated on serine residues by wheat germ Ca(2+)-dependent protein kinase (CDPK). [32P]phosphoWBP exactly comigrates with WBP on SDS-PAGE. WBP does not inhibit either wheat germ CDPK or calmodulin-dependent myosin light chain kinase. Apart from histone H1, WBP is the best endogenous substrate yet found for wheat embryo CDPK. A 12 kDa pine basic protein (PBP) was purified to homogeneity from seeds of stone pine (Pinus pinea L.) by a simple procedure involving batchwise elution from carboxymethylcellulose and cation exchange HPLC. PBP is also a good substrate for CDPK and is phosphorylated on Ser residues. N-terminal sequencing of WBP and PBP revealed that these proteins are homologous to a family of small basic plant proteins having a phospholipid transfer function.
Polya
G M
GM
Department of Biochemistry, La Trobe University, Bundoora, Victoria, Australia.
Chandra
S
S
Chung
R
R
Neumann
G M
GM
Höj
P B
PB
D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000737:chemistry / Q000302:isolation & purification*; D008565:Membrane Proteins / Q000737:chemistry / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D049294:Phospholipid Transfer Proteins; D010766:Phosphorylation; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification*; D011494:Protein Kinases / Q000737:chemistry*; D012639:Seeds / Q000737:chemistry / Q000201:enzymology; D012689:Sequence Homology, Nucleic Acid; D013379:Substrate Specificity; D014908:Triticum / Q000737:chemistry* / Q000196:embryology / Q000201:enzymology; D017871:Calcium-Calmodulin-Dependent Protein Kinases
D002352:Carrier Proteins; D008565:Membrane Proteins; D049294:Phospholipid Transfer Proteins; D010940:Plant Proteins; D011494:Protein Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases