Similar to its molecular relative hemoglobin, which manages the O₂ transport from the lung to the cells, myoglobin reversibly binds O₂ but with a sixfold higher affinity than the – with the bloodstream circulating – hemoglobin. Thereby, it can easily take over the hemoglobin-provided O₂ from the capillaries. O₂ binds to the central iron atom of the heme group (marked in orange at the left; also compare the section about ¹H NMR).

For the oxymyoglobin so formed three different functions were up to now suggested, which were mainly derived from in vitro experiments with pharmacological inhibition of myoglobin:

• Storage of O₂ as emergency reservoir
• Facilitated O₂ diffusion, that means in parallel to the diffusion of physically dissolved O₂, a transport of myoglobin-bound O₂ takes place from the capillary to the mitochondria
• Myoglobin-mediated oxidative phosphorylation

While the function of myoglobin as O₂ reservoir is generally accepted – which is in particular supported by the up to tenfold higher myoglobin content in muscles of diving vertebrates, such as dolphins and whales – both of the other functions were under dispute for decades due to contradictory data in the literature. The entire research area was revived some years ago by the generation of myoglobin-deficient mice (see figure top right) in our and an american laboratory. By comparison of normal control animals with the myoglobin lacking mutant we could show, that the facilitated O₂ diffusion by myoglobin indeed plays a major role for the adequate O₂ supply of the heart, and that on the other hand the myoglobin-mediated oxidative phosphorylation is only, if at all, of less significance (Merx et al, 2001).

What's more, we could discover surprisingly two important novel functions of myoglobin in muscle metabolism: The ability of myoglobin to effectively degrade both nitric oxide (NO) as well as free reactive oxygen species (compare also the original publications given below). The key to all of these investigations was the noninvasive measurement of myoglobin in the heart by ¹H NMR spectroscopy (see next section).

When the heart tissue is adequately supplied with O₂, myoglobin – owing to its high O₂ affinity – is completely oxygenated (MbO₂), and the methyl protons of the valine can be detected at a chemical shift of -2.7 ppm in the ¹H NMR spectrum of the heart (bottom trace). Only at a mismatch between O₂ supply and consumption myoglobin gets deoxygenated which is reflected in an intensity decrease of the MbO₂ signal. In that, the magnitude of this signal permits an assessment of the myocardial O₂ supply. Making use of a simple mathematical transformation the intensity of MbO₂ signal can be utilized to calculate the intracellular O₂ partial pressure (PO₂) of the heart muscle cells – a quantity, which is very difficult to access with other methods.

Taken together, monitoring the protons of the valine 68 by ¹H NMR spectroscopy as reporter nuclei of the electronic situation at the heme group yields versatile information about oxygenation, ligands, and redox state of cardiac myoglobin, and thereby about the current metabolic state of the heart.

At [NO] = 5 µM approximately 50% of the MbO₂ was converted to metMb reaching nearly 100% conversion at 25 µM NO (see also the quantitative analysis of this experiment). This reaction was reversible after cessation of the NO infusion (top trace), reflecting the rapid regeneration of MbO₂ by metMb reductase activity (compare the scheme below). It is noteworthy that the conversion of MbO₂ to metMb is already detectable at NO concentrations at which ³¹P NMR spectra did not yet show any adverse effect on cardiac energy status (third trace from top).

Of fundamental importance for the efficacy of this hoover is the activity of the endogenous metMb reductase which ensures the regeneration of Mb from metMb. Subsequent association with O₂ leads to reformation of MbO₂ available for another NO degradation cycle. This "molecular firewall" works highly efficient, so that even a massively raised NO formation by an overexpression of the inducible NO synthase (iNOS) can suprisingly be neutralized.