Evidence That Barley 3-Hydroxy-3-Methylglutaryl-Coenzyme A Reductase Kinase Is a Member of the Sucrose Nonfermenting-1-Related Protein Kinase Family

Abstract

A protein kinase was partially purified from barley (Hordeum vulgare L. cv Sundance) endosperm by ammonium sulfate fractionation, followed by ion-exchange, Reactive Blue, Mono-Q, and phosphocellulose chromatography. It was shown to phosphorylate Arabidopsis 3-hydroxy-3-methylglutaryl-coenzyme A (HMC-COA) reductase and a synthetic peptide that was shown previously to act as a substrate for HMC-COA reductase kinase purified from cauliflower, confirming it to be barley HMG-COA reductase kinase. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the partially purified preparation showed the presence of a polypeptide with an approximate relative molecular weight (M,) of 60,000, which is the size predicted for the barley sucrose nonfermenting-1 (SNF1)-related protein kinases BKIN2 and BKIN12. Antisera were raised to a rye (Secale cereale 1.) SNF1-related protein kinase (RKIN1) expressed in Fscherichia coli as a fusion with maltosebinding protein and to a synthetic peptide with a sequence that is conserved in, and specific to, plant members of the SNF1-related protein kinase family. l h e maltose-binding protein-RKIN1 fusion protein antiserum recognized a doublet of polypeptides with an approximate M, of 60,000 in crude endosperm extracts and a single polypeptide in root extracts, which co-migrated with the smaller polypeptide in the endosperm doublet. Both antisera recognized a polypeptide with an approximate M, of 60,000 i n the partially purified protein kinase preparation, suggesting strongly that barley HMC-COA reductase kinase is a member of the SNFl-related protein kinase family.

Biochemical and Functional Characterization of PKS11, a Novel Arabidopsis Protein Kinase

“…For example, the K m and V max values of a partially purified barley SnRK1 kinase, 3-hydroxy-3-methylglutaryl-coenzyme A reductase kinase, for a synthetic peptide SAMS (HMRSAMSGLHLVKRR), were determined to be 47 M and 25 nmol/min/mg, respectively (46). The apparent K m for ATP of a spinach SnRK1 was ϳ6 M (21).…”

Phosphoproteomic identification of targets of the Arabidopsis sucrose nonfermenting-like kinase SnRK2.8 reveals a connection to metabolic processes

“…Once a 14-3-3 binds to NR, the phosphoserine-NR is inactive (34). Many enzymes in C metabolism, such as sucrose phosphate synthase, sucrose synthase, and 3-hydroxy-3-methylglutaryl CoA reductase (8,32,35), are also controlled by phosphorylation and the binding of 14-3-3. The targets of most of the plant 14-3-3 have not been identified, and many of the plant 14-3-3 are poorly characterized (36).…”

“…SnRK1 kinases in plants have been well characterized at molecular and biochemical levels (8). In potato, SnRK1 is necessary for the sucrose-induced expression of sucrose synthase (9).…”

Signal Perception and Transduction: The Origin of the Phenotype.

“…Such disturbances may be detected by sucrose nonfermenting-1 (SNF-1) kinases, the activities of which are regulated by the cellular AMP/ATP ratio (Barker et al, 1996). Organic solvents are likely to increase plasma membrane permeability.…”