Date of Award:
5-1-1992
Document Type:
Dissertation
Degree Name:
Doctor of Philosophy (PhD)
Department:
Biology
Department name when degree awarded
Biology/Molecular Biology
Committee Chair(s)
Jon Y. Takemoto
Committee
Jon Y. Takemoto
Committee
Anne J. Anderson
Committee
Thomas A. Grover
Committee
Richard J. Mueller
Committee
Raghubir P. Sharma
Abstract
The syringomycin stimulated protein phosphorylation of red beet plasma membranes was investigated. Rabbit antisera directed against N- and C-termini and the nucleotide binding site of the H+-ATPase detected the 100 kD polypeptide of this enzyme in the plasma membrane. Immunoprecipitation experiments revealed that the 100 kD polypeptide was phosphorylated, and syringomycin stimulated the phosphorylation in a Ca2+-dependent manner. Phosphoamino acids in the immunoprecipitates were phosphoserine and phosphothreonine. These findings are evidence that syringomycin stimulates the phosphorylation of the plasma membrane H+-ATPase. Okadaic acid, a phosphatase inhibitor, did not affect protein phosphorylation of syringomycin-treated and KI-extracted plasma membranes. Therefore, protein kinases, but not protein phosphatases, are involved in the syringomycin effect. The plasma membrane was extracted with acetone and fractionated into three fractions (fractions A, B, and C). Syringomycin did not influence the protein kinase activities of the fractions, but Ca2+ stimulated fraction B activity. Thus, syringomycin does not directly stimulate protein kinases. Two protein kinases (MPKI and MPKII) were resolved by non-denaturing PAGE of fraction B, and were isolated by Cibacron-Blue agarose chromatography. Both kinases were Ca2+-dependent, capable of phosphorylating casein, and were not affected by syringomycin, cAMP, or phospholipids. MPKII was highly sensitive to the calmodulin antagonist, trifluoperazine. MPKI was purified to homogeneity using ATP-agarose chromatography. Its major 58 kD polypeptide subunit has an N-terminal amino acid sequence of K,W/G/S,L/N,E,E,L,N,I/V,Q,G. MPKI is a novel plant plasma membrane protein kinase, which, together with MPKII, is possibly involved in the regulatory phosphorylation of the H+-ATPase.
Recommended Citation
Suzuki, Yuka S., "Red Beet Ca2+-Dependent Protein Kinases and Their Role in Syringomycin Action" (1992). Biology. 589.
https://digitalcommons.usu.edu/etd_biology/589
Copyright for this work is retained by the student. If you have any questions regarding the inclusion of this work in the Digital Commons, please email us at .