Notable diversity in hemoglobin expression patterns among species of the deep-sea clam, Calyptogena

The deep-sea clams Calyptogena nautilei and C tsubasa, which live in the cold-seep area at a depth of 3570 m in the Nankai Trough, Japan, have abundant hemoglobins (Hbs) in erythrocytes, similar to other Calyptogena species. We determined the cDNA-derived amino acid sequences of Hbs from two Calyptogena species. C tsubasa was found to contain two dimeric Hbs, Hb I consisting of 145 amino acid residues and Hb II with 137 residues, similar to known Hbs from C soyoae and C kaikoi. Sequence identity was over 90% among the orthologous chains of Calyptogena Hbs. On the other hand, surprisingly, C nautilei contained two monomeric Hbs, Hb III containing 141 residues and Hb IV with 134 residues. In addition, Hbs III and IV showed only 33-42% sequence identity with Hbs I and II from other Calyptogena species. The distal (E7) histidine, one of the functionally important residues of the heme protein, is replaced by glutamine in all Hb chains of Calyptogena species. A phylogenetic analysis indicated that C nautilei Hb III is closer to Hb I from other Calyptogena species. We suppose that a Hb gene was duplicated at least three times in an immediate ancestor of Calyptogena and, presumably depending on physiological conditions different Hb sets are being expressed: dimeric Hbs I and II in C soyoae, C kaikoi and C tsubasa, and monomeric Hbs III and IV in C nautilei.