title: Structural effects of protein lipidation as revealed by LysB29-myristoyl, des(B30) insulin.
abstract: Intracellular proteins are frequently modified by covalent addition of lipid moieties such as myristate. Although a functional role of protein lipidation is implicated in diverse biological processes, only a few examples exist where the structural basis for the phenomena is known. We employ the insulin 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 the modification. However, the flexible positions taken up by the hydrocarbon chain selectively modify key structural properties. In the insulin monomer, 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% increase in the free energy of unfolding of the protein. Addition of two Zn(2+)/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), T(3)R(3), and R(6) allosteric states of the insulin hexamer. We conclude that insulin is an attractive model system for analyzing the diverse structural effects induced by lipidation of a compact globular protein.
authors.affiliation: Health Care Discovery, Novo Nordisk A/S, Novo Alle 6B1, DK-2880 Bagsvaerd, Denmark.
authors.forename: H B
authors.initials: HB
authors.lastname: Olsen
authors.forename: N C
authors.initials: NC
authors.lastname: Kaarsholm
mesh_terms: D000215:Acylation; D000817:Anilino Naphthalenesulfonates / Q000737:chemistry; D002352:Carrier Proteins / Q000737:chemistry; D002942:Circular Dichroism; D019281:Dimerization; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D007328:Insulin / Q000031:analogs & derivatives* / Q000737:chemistry*; D000069057:Insulin Detemir; D049528:Insulin, Long-Acting; D008239:Lysine / Q000737:chemistry*; D019814:Myristic Acid / Q000737:chemistry*; D019906:Nuclear Magnetic Resonance, Biomolecular; D010446:Peptide Fragments / Q000737:chemistry*; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D013056:Spectrophotometry, Ultraviolet; D013816:Thermodynamics
chemical_list: D000817:Anilino Naphthalenesulfonates; D002352:Carrier Proteins; D005456:Fluorescent Dyes; D007328:Insulin; D049528:Insulin, Long-Acting; D010446:Peptide Fragments; C414550:insulin, Lys(B29)-(N(epsilon)-myristoyl)-des(b30); D019814:Myristic Acid; D000069057:Insulin Detemir; C027132:1-anilino-8-naphthalenesulfonate; D008239:Lysine