Kinetic studies of the yeast His-Asp phosphorelay signaling pathway.
For both prokaryotic and eukaryotic His-Asp phosphorelay signaling pathways, the rates of protein phosphorylation and dephosphorylation determine the stimulus-to-response time frame. Thus, kinetic studies of phosphoryl group transfer between signaling partners are important for gaining a full understanding of how the system is regulated. In many cases, the phosphotransfer reactions are too fast for rates to be determined by manual experimentation. Rapid quench flow techniques thus provide a powerful method for studying rapid reactions that occur in the millisecond time frame. In this chapter, we describe experimental design and procedures for kinetic characterization of the yeast SLN1-YPD1-SSK1 osmoregulatory phosphorelay system using a rapid quench flow kinetic instrument.
Kaserer
Alla O
AO
Department of Chemistry and Biochemistry, University of Oklahoma, Norman, Oklahoma, USA.
Andi
Babak
B
Cook
Paul F
PF
West
Ann H
AH
D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology*
D047908:Intracellular Signaling Peptides and Proteins; C087199:SSK1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D011494:Protein Kinases; C102035:YPD1 protein, S cerevisiae; C083749:SLN1 protein, S cerevisiae