Natural abundance 13C spectra of proteins: Carboxy-myoglobin and hemoglobin

Abstract

In the past few years the high resolution NMR technique has been successfully applied in studies of biologically important molecules. However, due to the large number of the protons present, the proton NMR spectra of the proteins studied [l-3] are so poorly resolved that a detailed complete interpretation has never been possible. Despite this complexity, the assignments of some resonances to particular groups was possible, when their relative changes furnished some indications of the conformational variations occurring. This is so of the proton resonances of the amino acid residues near to the heme group in hemoglobin and myoglobin (i.e. histidine residues) which are differently affected in the oxy and deoxy forms by the ring current, contact, and pseudocontact shifts [3]. However, no detailed and complete analysis of the spectra has been achieved. The relative simplicity of the 13C natural abundance spectra resulting from large chemical shifts, absence of any spin-spin coupling, relatively narrow lines not seriously broadened by dipolar interactions, suggests that better resolution and more detailed information oan be obtained from 13C NMR spectra. The chief obstacle to date has been the low sensitivity resulting from the character of the ’ 3C nucleus itself and from its low natural abundance (1 per cent) [4,8]. This limitation has been overcome by the use of the Fourier transform NMR technique [9, lo], In this paper we report the results obtained with this technique for myoglobin and hemoglobin.