1H n.m.r. studies of insulin. Reversible transformation of 2-zinc to 4-zinc insulin hexamer.

1H n.m.r. studies at 270 MHz were made of the transformation of 2 Zn insulin hexamer to 4 Zn hexamer produced by the addition of anions (thiocyanate ion). Four separate H2 histidine resonances were observed for the B5 and B10 histidines in 2 Zn hexamer at pH 7 and 9 and four separate resonances also occurred in the 4 Zn hexamer. The observation of these resonances and others from phenylalanine, tyrosine and leucine residues showed that the 2 Zn to 4 Zn transformation probably occurred in solution in a similar manner to that observed in the crystal. Furthermore as occurred in the crystal, it was found that in solution the transformation was reversible (on removal of thiocyanate) and that 2 Cd insulin was unable to undergo the transformation. Des-Phe-Bl-insulin did not undergo the transformation. Addition of SCN- to Zn-free insulin (mainly dimer) produced only a small transformation, consistent with the idea that Zn2+ promotes formation of hexamer from dimer but probably does not otherwise affect the transformation.