Date of Award


Document Type


Degree Name

Master of Science (MS)



First Advisor

Margaret Vizzard


Cyclic GMP-dependent protein kinases (PKG’s) are essential signaling macromolecules which play a pivotal role in vascular physiology and smooth muscle tone regulation. As principle downstream effectors of the secondary messenger cyclic 3’, 5’-guanosine-monophosphate (cGMP), PKG isoforms are expressed in high levels in all types of smooth muscle cells. The broad range of cellular functions effected by PKG include platelet aggregation, hypertrophy, apoptosis, neuronal plasticity, gene expression, differentiation, vasorelaxation, vascular remodeling, calcium homeostasis, and cardiac function. Recently, a newly characterized helical switch domain within the alpha isoform of PKG (PKG1-α) has led to the development of S1.1, a novel cGMP-independent peptide activator of PKG1-α. The ability to activate PKG1-α independent of cGMP could have great significance in the development of therapeutics which target vascular smooth muscle. The molecular mechanism behind S1.1 activation of PKG1-α remains unclear. However, a series of S1.1 derivatives have begun to elucidate the synthetic peptide (S-tide) pharmacophore. Until now, the canonical theory of the S-tide pharmacophore involved hydrophobic interactions provided by two phenylalanine residues within the S-tides that are analogous to a portion of the parent enzyme which has been labeled the knob motif. This hypothesis was largely based on the results of previous experiments which showed a complete loss of activity with S1.6, a derivative lacking the phenylalanine residues.The work displayed in this thesis aims to question the current theory of the synthetic peptide pharmacophore and instead suggest that the loss of activity observed with S1.6 was a consequence of the radioactive kinase assay that was being used to measure activity. By using a slight deviation in methodology, it has been shown that it is possible to drastically alter S-tide activation kinetics. The implications of this are that the apparent S-tide kinetic profile has varied wildly across studies based on which methodology was being employed. The assay component responsible for this shift in activity was isolated through iterative deconstruction of the assay and determined to be magnesium acetate (MgOAc). How MgOAc modulates S-tide activation remains to be discovered, however, these findings are significant and require all past S-tide studies to be examined in a critical light based on these results.



Number of Pages

68 p.