Genetic and biochemical studies of a mutant Saccharomyces cerevisiae myristoyl-CoA:protein N-myristoyltransferase, nmt72p super(Leu99 arrow right Pro), that produces temperature-sensitive myristic acid auxotrophy

Saccharomyces cerevisiae myristoyl-CoA:protein N-myristoyltransferase (nmt1p) is an essential enzyme that transfers myristate from CoA to the amino-terminal glycine residue of at least 12 cellular proteins. Its reaction mechanism is Ordered Bi Bi with myristoyl-CoA binding occurring before binding of nascent polypeptides and release of CoA preceding release of the myristoylprotein product. nmt1-72 is a temperature-sensitive allele, identified by Stone et al. that causes arrest in the G1 phase of the cell cycle due to reduced acylation of Gpa1p. We have recovered this mutant allele and determined that it contains a single point mutation resulting in a Leu super(99) (CTA) to Pro (CCA) substitution. Addition of greater than or equal to 500 mu M myristate but not palmitate to synthetic or rich media rescues the growth arrest caused by nmt1-72 at 37-39 degree C, consistent with the observation that purified nmt72p has reduced affinity for myristoyl-CoA and that exogenous myristate but not palmitate increases cellular myristoyl-CoA pools.