Hemocyanin Octopus

Figure 7 of reference 11 compares similarities between Limulus and Octopus hemocyanins by illustrating the overlap of 30 amino acids about the CuB region. The authors find that all residues within 5.0 A of the Cu-Cu midpoint are conserved between arthropodan and molluscan hemocyanins and occupy analogous positions except for one (see the following paragraph). The three histidines about CuB overlap almost exactly in orientation and are less than 1 A apart. The histidines... 

In a study of octopus hemocyanin (Salvato et al., 1989), a green half-methemocyanin could be formed in the presence of slight molar excess of sodium nitrite and ascorbate. This form was believed to consist of cuprous and cupric metals, but the amoung of NO directly bound was less than 0.1 mol. NOi was believed to be the oxidizing species its relationship to the endogenous ligand was not addressed. Apparently, hemocyanins are most likely to be found in deoxy, or reduced, forms, and it is difficult to oxidize them. 

Molluscan hemocyanins. Two FUs from moUuscan hemocyanins were resolved, the oxy-form of O. dofleini He FU g (Figure 5b) and the deoxy-form of R. thomasiana He (Figure 5c ). Each FU consists of two domains. The N-terminal domain II carries the active site with a four alpha-helix bundle folding motif with two copper atoms. The C-terminal domain III replaces topologically the domain I in arthropod subunits and looks like a squeezed beta-barrel. Although the Rapana structure is not resolved as well as the Octopus FU, two different conformations can be deduced. In the oxy FU of Octopus hemocyanin, domain III covers the entrance to the active site completely while in the deoxy-form this domain is shifted a few degrees so that the channel to the active site becomes completely uncovered. 

Ipomoea catecholoxidase, corresponds to Leu2830 in Octopus hemocyanin, which blocks the entrance. 

Figure 3 Binuclear copper complexes in octopus hemocyanin (a) and sweet potato catechol oxidase (b) showing the thioether linkage between a cysteine residue and a directly coordinating histidine imidazole. Heteroatoms (N,0,S) are indicated by a shaded quadrant. [ortep-III views based on PDB ID 1JS8 and IBTl]...

Figure 3. Oscilloscope trace of a temperature-jump experiment on Octopus hemocyanin reacting with oxygen. Potassium phosphate buffer, 0.2M, pH 7, and 20 C (before the jump). Discharge 30 kv yielding a temperature increase of 4 to 5 C. Protein concentration = 4.5 X 10 binding equivalent L fractional saturation with oxygen = 0.53 free oxygen concentration = 3.4 X lOr M. Sweep time = 100 fxsec per large screen division observation wavelength = 348 nm (24).

In 1847 E. Harless discovered the presence of copper in the blood of the octopus Eledone and the snail Helix pomatia (172, 173). Investigation of the phenomenon by which the blood and tissues of certain marine animals turn blue on exposure to air finally led to the discovery that the blood plasma of such animals contains copper combined with a protein. Because of its analogy to hemoglobin and its ability to carry oxygen, L. Fredericq in 1878 named the copper-containing protein in the blood of Octopus vulgaris hemocyanin (173, 174). 

L. B. Mendel and H. C. Bradley found in 1905 diat the snail syco-typus contains zinc in the fiver and in die oxygen-carrying protein of the blood, liemosycotypin. The diree respiratory proteins, hemoglobin of the vertebrates, hemocyanin of die octopus, and hemosycotypin of the... 

Figure 2 Active sites of hemocyanins from horseshoe crab L. polyphemus (a oxy b deoxy), spiny lobster Panulirus interruptus (c), sweet potato Ipomoea batatas (d), Octopus dofleini (e), Rapana thomasiana (f). Cleary the four alpha-helix bundle motif with the metal center can be seen. The three histidines coordinating Cu-A are colored red, those coordinating Cu-B green. The two copper atoms are colored blue and oxygen red. The cysteines binding covalently a histidine at the Cu-A site are colored yellow. In the case of Ipomoea, a water molecule connects the two copper atoms...

Although the amino acid sequence of tyrosinase has only 25.3 and 26.0% identities with those of the /. batatas catechol oxidase and the odg domain of the Octopus dofleini hemocyanin, " respectively, its overall structure is quite similar to theirs. Among these three proteins, a high degree of conservation is observed in the core domain composed of the ct-bundle. The tyrosinase and hemocyanins from Panulirus interruputus and... 

Alternative reaction mechanisms include a radical mechanism proposed by Kitajima and Morooka and a mechanism involving a Cu(III) intermediate based on measurements of model compounds. On the basis of the crystal structure of the catechol oxidase-PTU inhibitor complex, monodentate binding of the substrate was suggested for catechol oxidase. A radical mechanism, as proposed for the weak catecholase activity found in Octopus vulgaris hemocyanin, is also possible for catechol oxidase due to the strong structural relationship between catechol oxidase from /. batatas and odg hemocyanin as described above. 

Unlike mammals, snails, spiders, and octopuses do not use hemoglobin to transport oxygen. Instead, they rely on a related compound known as hemocyanin. This molecule doesn t have an atom of iron in its middle it has an atom of copper, which binds oxygen. Hemocyanin absorbs all colors except blue, which it reflects. 

In certain copper-containing proteins the copper appears to serve principally in electron transport with no evidence of CU-O2 interaction, such as in cytochrome oxidase. Of importance, however, is that many copper proteins and enzymes participate in reactions in which the oxygen molecule is directly or indirectly involved. An example is hemocyanin, the oxygen carrier in the blood of certain sea animals such as snails, octopus, and Crustacea. Oxygenated hemocyanin is blue and the cephalopods (crabs and lobsters) are literally the blue bloods of the animal kingdom. Hemocyanins are giant molecules of MW > 10 that occur free in solution. 

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