Abstract
The myoglobins from the sharks Galeorhinus japonicus and Musterus japonicus possess a distal Gln-E7 instead of the usually found His-E7. The met-cyano form of these shark myoglobins has been studied by 1H- and 15N-NMR in order to gain insight into the functional properties of the Gln-E7. The analysis of paramagnetic relaxation has provided the assignment of the resonance arising from one of the Gln-E7 N ϵH labile protons, whilst the rate of its chemical exchange has been analyzed in detail by using a saturation transfer method. The hydrogen-bonding interaction between this proton and Fe-bound-CN − has been clearly manifested in the hyperfine shift of the Gln-E7 N ϵH proton resonance as well as its chemical exchange behavior. The resonances of the Gln-E7 side-chain non-labile protons have been partly assigned on a basis of both scalar and dipolar connectivities. The analysis of the dipolar connectivities among the side-chain protons and the iron-proton distances determined from their paramagnetic relaxation rate has revealed that the side chain adopts a conformation with its carbonyl oxygen oriented away from the heme. Although 1H-NMR spectra of these two myoglobins are essentially similar, a relatively large difference in the shift of Gln-E7 N ϵH proton and Fe-bound C 15N − resonances between the two has been observed which is attributed to a differential hydrogen-bonding interaction between these proteins. The present study demonstrates the sensitivity of NMR parameters to the hydrogen-bonding interaction between coordinated ligand and a distal amino-acid side chain in paramagnetic hemoproteins.
Published Version
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