A Conformational Study of Glucagon

Abstract

glucagon is a polypetide of 29 residues, which can be crystallised. Optical rotatory dispersion in dilute neutral, acid and alkaline solution suggests that the polypeptide chain is largely random, but contains about one turn of α‐helix, which is eliminated in 6 M guanidine hydrochloride. Using thermal difference spectra, nuclear magnetic resonance, and the temperature‐dependence of the optical retatory dispersion as criteria, it appears that glucagon possesses no difined tertiary structure in solution, but may be regarded as being in a state of α‐helix ⇌ random coik equilibrium, near the high‐temperature end of the transition range. That there is no steric objection to the formation of a highly α‐helical conformation is demonstrated by optical rotatory dispersion measurements in 2‐chloroethanol solution, in which the protein becomes largely helical. A rough calculation indicates that in aqueous solution at pH 2 the transition mid‐point is at about –40%%. From changes in the proton magnetic resonance spectrum when glucagon is transferred from an aqueous environment into 6 M guanidine hydrochloride, it is deduced that the short helical segment in the chain in the aqueous state contains most of the long aliphatic side chains, and since the sequence is known it has been possible to identify such as segment uniquely near the C‐terminal end. On standing in acid solution the vicosity increases and a birefringent gel is formed. Sedimentation studies indicate the formation of large aggregates. On further standingor warming, a precipitate appears which has the appearance of long fibrils in the electron microscope. Infrared spectra of the gel, of solid films and of the precipitated material show that in all these states the glucagon is in the form of antiparallel β‐chains.