P27 is a tumor-suppressor protein involved in cell cycle regulation. It functions by binding and inhibiting activity of cyclin-dependent kinases (CDKs). Surprisingly, the p27 gene is seldom mutated or deleted. Instead, p27 is deregulated at the protein level such that its tumor suppressing ability is compromised, thereby contributing to development of many types of aggressive cancer. For this reason understanding the molecular mechanisms controlling p27 function is essential. The p27 protein is unique in that it lacks significant tertiary structure and is heat stable. Thus, we hypothesized that compromising p27 function might involve imparting structure to the polypeptide. A search for enzymes having the ability to alter protein structure identified transglutaminase, which catalyzes protein cross-linking via formation of an isopeptide bond between lysine and glutamine groups. Interestingly, altered levels of nuclear and cellular transglutaminase have been observed in cancer cells. We therefore incubated purified p27 and transglutaminase, followed by analysis of p27 mobility on a denaturing polyacrylamide gel. Surprisingly, p27 monomers were not crosslinked, Instead, p27 migrated faster, suggesting a conformational change due to intramolecular isopeptide bond formation. The kinetics, inhibition, and specificity of this novel reaction have been evaluated. Our results identify a possible link between transglutaminase modification of P27 and proliferation of aggressive cancers.

Modification of the p27 tumor suppressor by the transglutaminase enzyme

249th National Meeting (2015)

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