Sequence and evolutionary studies on hemoglobin from the sea cucumber Molpadia arenicola

The amino acid sequence of the coelomic hemoglobin Hb C from the sea cucumber Molpadia arenicola was determined. Samples of C globin were digested with various proteolytic enzymes and chemical reagents. Peptides generated by these digests were separated and purified by high voltage paper electrophoresis and high performance liquid chromatography. The sequences of these peptides were determined by manual or automated Edman sequencing. Overlapping peptide sequences were assembled to determine the amino acid sequence of Hb C.;The sequence of Hb C was examined for homology to the previously determined sequence of Molpadia arenicola Hb D. The two globins were found to be 59% identical in sequence. On the basis of this large degree of identity, it was determined that the divergence of the coelomic globins of Molpadia arenicola was a more recent event than the divergence of the echinoderms from the vertebrates. Comparison of Hb C to other vertebrate and invertebrate globin sequences indicated no strong homology to known globins from any other phyla. Hb C was found not to greatly resemble either vertebrate alpha-like or beta-like hemoglobins. This, together with the lack of alpha-like or beta-like character of Hb D, led to the conclusion that the globins of Molpadia arenicola do not function as a globin system similar to vertebrate alpha and beta globins.;Partial sequence analysis of a third Molpadia arenicola globin, Hb A, was also completed. The regions of Hb A sequenced were found to be quite similar to Hb C, as had been indicated by earlier immunological data.;The level of glycosylated hemoglobin in the coelomic, intestinal and water vascular fluid of Molpadia arenicola was determined by phenylboronate affinity column chromatography. Glycosylated hemoglobin levels were found to be similar to vertebrate levels in all three Molpadia systems, as well as in the invertebrates Barbatia reeveana and Urechis caupo. However, the globins from Lumbricus terrestris and Cardita affinis were found to be more highly glycosylated.