Complete amino-acid sequence of a functional unit from a molluscan hemocyanin (Helix pomatia).
From the beta c-hemocyanin (beta c-Hc) of the vineyard snail, Helix pomatia, the functional unit d (Mr approximately equal to 50,000-55,000) was isolated by limited proteolysis and gel chromatography. A small quantity of functional unit d was obtained intact, but the major part in the form of two peptides (Mr approximately equal to 43,000 and 10,000, respectively) connected by a disulfide bridge. After reduction and carboxymethylation, these were separated from each other and cleaved by conventional methods. The peptides were isolated by gel chromatography and HPLC, and sequenced manually or automatically. The complete sequence of Helix beta c-Hc d comprises 410 residues plus 3 residues at the N-terminus seemingly resulting from incomplete cleavage. There is apparently only one carbohydrate side-chain. Comparison of this gastropodan hemocyanin sequence to the partial sequence of a cephalopodan Hc C-terminal unit revealed sufficient identities to state that the functional units of molluscan hemocyanins have arisen by a series of gene duplications. On the other hand, there is practically no homology with arthropodan hemocyanins except for one section of 42 residues which is clearly homologous. This section corresponds to the "Copper B" site of Panulirus interruptus hemocyanin. It is also found in tyrosinases from Neurospora crassa, Streptomyces glaucescens, and mouse. In the N-terminal half of Helix beta c-Hc d there are other sections clearly homologous to the tyrosinases, but overall homology is limited. The second copper-binding site was not identified but must be completely distinct from the "Copper A" binding site of arthropodan hemocyanins. It is suggested that molluscan and arthropodan hemocyanins have evolved independently from a common ancestral mononuclear copper protein.
D000595:Amino Acid Sequence; D000596:Amino Acids; D000818:Animals; D002850:Chromatography, Gel; D003488:Cyanogen Bromide; D004591:Electrophoresis, Polyacrylamide Gel; D006372:Helix, Snails / Q000378:metabolism*; D006433:Hemocyanins / Q000032:analysis*; D006868:Hydrolysis; D011487:Protein Conformation; D014357:Trypsin
D000596:Amino Acids; D006433:Hemocyanins; D014357:Trypsin; D003488:Cyanogen Bromide