Protein N-myristoylation: Novel target for antitumor and antiviral agents. Characterization of analogs of myristoyl-CoA as inhibitors of myristoyl-CoA:protein N-myristoyltransferase

Lisa A Paige, Purdue University

Abstract

The covalent attachment of the 14-carbon saturated fatty acid, myristic acid, to the amino-terminus of certain viral and cellular proteins has important implications for both the localization and function of the modified protein. The critical role of myristoylation for the oncogenic potential of the protein-tyrosine kinase pp60$\sp{v-src}$ and for the ability of retroviral gag proteins to participate in viral assembly, has engendered interest in the design of inhibitors of myristoyl-CoA:protein N-myristoyltransferase (NMT), the enzyme which catalyzes myristoylation. We have shown that analogs of myristoyl-CoA can serve as effective inhibitors of NMT both in vitro and in cultured cells. S-(2-oxopentadecyl)-CoA, a nonhydrolyzable analog of myristoyl-CoA, is a potent in vitro inhibitor of NMT, exhibiting a K$\sb{\rm i}$ value of 24 nM. Additionally, myristic acid analogs with substitutions at the 2-position can become metabolically activated to form potent inhibitors of NMT in intact cells. 2-Hydroxy-, 2-fluoro- and 2-bromomyristoyl-CoA exhibited K$\sb{\rm i}$ values of 45,200 and 450 nM, respectively. 2-Fluoromyristoyl-CoA was the only one of these compounds which served as a substrate for NMT; however, the rate of transfer of 2-fluoromyristate to a peptide substrate was approximately 20-fold lower than that for myristate. Modifications were made to the potent inhibitor S-(2-oxopentadecyl)-CoA to probe the structural features which were required for binding by NMT. Most notably, removal of the adenosine phosphate moiety, or reduction of the acyl-carbonyl on this substrate analog resulted in a complete loss of binding by NMT. We have also made the observation that the myristoylated lymphocyte specific protein-tyrosine kinase p56$\sp{lck}$ is also acylated with palmitic acid. This modification may play an important role in regulating the interactions of p56$\sp{lck}$ with other proteins involved in the activation of T lymphocytes. Investigations aimed at identifying the amino acid residue modified by palmitate and determining the role of this modification are currently underway.

Degree

Ph.D.

Advisors

Geahlen, Purdue University.

Subject Area

Biochemistry

COinS