JMS/sandbox15

Extraordinary ProteinsExtraordinary Proteins

Life has managed to weather extreme environments - almost every hole we've poked a stick into contains thriving living communities. Proteins are a necessity for living, and therefore tuning protein structures to an extreme environment is of paramount value to an evolving organism. In this article, we present the biophysical modifications present in extreme protein structures.

Positively charged myoglobin allows whales to hold their breath during long divesPositively charged myoglobin allows whales to hold their breath during long dives

Elephants can hold their breath for 2 minutes, but whales can hold their breath for 60 minutes - and they do, migrating underwater around the world. A group of researchers contacted museums and zoos around the world.

They collected muscle tissue from hundreds of aquatic and terrestrial mammalian species. myoglobin stores oxygen in muscle tissue, and the researchers thought they might find differences relating to this protein in terrestrial versus aquatic animals. They measured the cellular concentration and net charge of myoglobin from each species muscle tissue sample. To measure the net charge, they sequenced each specie's myoglobin gene and computationally modeled the net charge. for a representative sample of species they additionally directly measured the net charge of the different myoglobin protein via native gel electrophoresis.

Their hypothesis was that whales and other aquatic animals can hold their breath for so long because they can store more oxygen in their muscles by increasing the concentration of myoglobin in each muscle cells. Specifically, they predicted that species could increase the concentration of myoglobin by increasing its solubility through increasing the net charge, so that there would be repulsion between the myoglobin protein even at high concentrations, which would prevent aggregation and precipitation.

Amazingly, they found an association between an animals' ability to hold its breath, high concentrations of myoglobin in muscle tissue, and a larger positive net charge of myoglobin. Typically, purified terrestrial mammal's myoglobin has a solubility of 20 mg/g in an aqueous solution at neutral pH ([Sigma Aldrich]) which turns out to be the maximum level of myoglobin found in most terrestrial mammal's tissue. But whales and other aquatic mammals far exceed this solubility limit, e.g., whales have 70 mg/g. The way that they overcome the solubility constraint may be traced back to a modest increase in the net charge of myoglobin - from around +2 in terrestrial animals to around +4 in aquatic animals.

Molecular Tour The ability of increasing net charge to enable higher solubility is a known phenomenon, and this study is consistent with previous reports. The aquatic animals have increased their net charge in a variety of ways - different combinations of amino acids switches. We present one such manifestation of this overall trend, by comparing the elephant and whale myoglobin structures. Out of 27 divergent amino acids between whale and elephant's myoglobin - from a total of 153 amino acids - only 8 of these amino acids lead to changes of charge. These eight amino acids are shown for whales and elephants, . The whale amino acids, have an illustrative eletrostatic field drawn around the electrically charged atom in the residue. Next to the whale amino acid, the elephant residues are shown in yellow halos and labelled with the residue name. From studying the differences between these two proteins, it is clear that the whale protein has more areas with a positive electrostatic field. These positive electrostatic fields are scattered about the whale protein, and will repel any whale myoglobin, preventing the protein-protein interactions that lead to aggregation. Another way to say this, in the case of proteins , is that water binds to the hydrogens coming off a positively charged atom, or associates with the charged atom itself, thus screening the protein from another protein, which is further screened itself. This picture, while illustrative of the basic principles of how small electrostatic fields changes can greatly increase a protein's solubility, is not completely rigorous. In fact, among other considerations, one needs to consider neighboring residues to these divergent residues, in order to calculate the actual electrostatic field. It may be in some cases, that a negatively charged residues neigbors one of these additional positive residues, and from a distance relatively larger than the distance between the two oppositely charged residues, the electrostatic field is zero. Asp and Glu have a charge of -1, Arg and Lys have a charge of +1, His in the positions shown here - 12 and 116 (Table S2) - have a charge of about +0.5.

spinqualitylabelsall models myoglobin (PDB entry 1mbn) Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image

However, a 3-fold increase in concentration of myoglobin ought to result in a similar fold increase in max time of breath holding, and the researchers show that body mass also makes a critical contribution to an animal's ability to hold its breath, with the overall equation for the contribution of body mass and myoglobin net charge as follows:

log (maximum time underwater) = 0.223*log(body mass) + 0.972*log(myoglobin net charge) + 0.891

As Asian elephant's weight is ~3K Kg, and a sperm whale's weight is ~50K Kg, it is clear that the modest increase in net charge contributes about the same as the enormous difference in body mass to the maximum time underwater.