Structural effects of protein lipidation as revealed by Lys(B29)-myristoyl, des(B30) insulin

Intracellular proteins are frequently modified by covalent addition of lipi d moieties such as myristate, Although a functional role of protein lipidat ion is implicated in diverse biological processes, only a few examples exis t where the structural basis for the phenomena is known. We employ the insu lin molecule as a model to evaluate the detailed structural effects induced by myristoylation. Several Lines of investigation are used to characterize the solution properties of Lys(B29)(N-epsilon-myristoyl) des(B30) insulin. The structure of the polypeptide chains remains essentially unchanged by t he modification. However, the flexible positions taken up by the hydrocarbo n chain selectively modify key structural properties. In the insulin monome r, the myristoyl moiety binds in the dimer interface and modulates protein- protein recognition events involved in insulin dimer formation and receptor binding. Myristoylation also contributes stability expressed as an 30% inc rease in the free energy of unfolding of the protein. Addition of two Zn2+/ hexamer and phenol results in the displacement of the myristoyl moiety from the dimer interface and formation of stable R-6 hexamers similar to those formed by human insulin. However, in its new position on the surface of the hexamer, the fatty acid chain affects the equilibria of the phenol-induced interconversions between the T-6, T3R3, and R-6 allosteric states of the i nsulin hexamer. We conclude that insulin is an attractive model system for analyzing the diverse structural effects induced by lipidation of a compact globular protein.