The fundamental goal of this research program is to improve understanding of the molecular mechanisms underlying hormonal control in mammalian cells. The model system has been the regulation of glycogen metabolism, and in particular the role of phosphorylation in the control of glycogen synthase. This enzyme, which is regulated by several hormones, such as insulin, epinephrine and glucagon, is subject to complex multisite phosphorylation. Previous study of several protein kinases that phosphorylate glycogen synthase has led to a mechanism whereby certain enzymes, casein kinase I and glycogen synthase kinase 3, appear to recognize substrates only after a separate, primary phosphorylation. Thus, it is proposed that glycogen synthase is phosphorylated by a 'hierarchal' mechanism, requiring the sequential action of multiple kinases. Recent work has also been directed at the pathway of the initiation of glycogen synthesis which involves a specialized protein called glycogenin. This is an interesting potential locus for the control of glycogen accumulation. The proposed research falls into three closely related areas. The first is the direct extension of earlier work and focuses on the control of glycogen synthase.
Aim (i) will address structure-function correlations for the enzyme, with an emphasis on the relative roles of individual phosphorylation sites.
Aim (iii) will focus on testing the performance of various glycogen synthase mutants once introduced into mammalian cells. Phosphorylation site mutants will test the hierarchal phosphorylation hypothesis in cells. Insulin controls will also be examined. The second focus is on glycogenin and the initiation pathway.
Aim (ii) is directed at the mechanism of activation and possible control of glycogenin.
In aim (iii) expression of glycogenin in mammalian cells will be investigated. The third focus, aim (iv), is on the casein kinase I family of protein kinases. Main interests are in potential regulation and in understanding the molecular basis of substrate recognition.
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