Physicochemical studies of the rat liver and adipose fatty acid synthetases.

Abstract

M guanidinium chloride by the analytical ultracentrifuge also indicated that the enzyme consists of two subunits of the same size. The possibility that 6 M guanidinium chloride does not result in complete dissociation of the complex into its ultimate subunits is unlikely since the circular dichroism spectrum of the enzyme in this solvent indicated that the enzyme exists as random coils. Binding studies with [‘4C]acetyl-CoA and [‘4C]malonyl-CoA were consistent with these observations in that they indicated that the 4’-phosphopantetheine and the catalytic centers are associated only with the 220,000 molecular weight subunit. Electron microscopy studies of the rat liver enzyme indicate that the subunit consists of a linear structure 200 A long containing at least four lobes 50 A in diameter. A pseudotetrahedral set of images was also observed and it was proposed that these images resulted from the folding of the linear structures into the pseudotetrahedral form. These observations indicate that the multicatalytic activities of the synthetases and the acyl carrier protein are associated only with the two polypeptide chains. Active enzyme centrifugation studies indicated that the dimer is the only catalytically active form of the enzyme. No larger active form of the enzyme was detected in the presence of the substrates. Furthermore, the monomeric form of the enzyme is inactive. The requirement of the dimer for activity may result from the involvement of two prosthetic groups, or a catalytic center comprising the two polypeptide chains in the catalytic process, or both.