The first complete cDNA sequence of the hemocyanin from a bivalve, the protobranch Nucula nucleus.
By cDNA sequencing we have achieved the first, and complete, hemocyanin sequence of a bivalve (Nucula nucleus). This extracellular oxygen-binding protein consists of two immunologically distinguishable isoforms, here termed NnH1 and NnH2. They share a mean sequence identity of 61%, both contain a linear arrangement of eight paralogous, ca.50-kDa functional units (FUs a-h), and in both isoforms the C-terminal FU-h possesses an extension of ca. 100 amino acids. The cDNA of NnH1 comprises 11,090 bp, subdivided into a 5'utr of 75 bp, a 3'utr of 791 bp, and an open reading frame for a signal peptide of 19 amino acids plus a polypeptide of 3389 amino acids (Mr = 385 kDa). The cDNA of NnH2 comprises 10,849 bp, subdivided into a 5'utr of 47 bp, a 3'utr of 647 bp, and an open reading frame for a signal peptide of 16 amino acids plus a polypeptide of 3369 amino acids (Mr = 387 kDa). In contrast to other molluscan hemocyanins, which are highly glycosylated, the bivalve hemocyanin sequence exhibits only four potential N-glycosylation sites, and within both isoforms a peculiar indel is present, surrounding the highly conserved copper-binding site CuA. Phylogenetic analyses of NnH1 and NnH2, compared to the known hemocyanin sequences of gastropods and cephalopods, reveal a statistically sound closer relationship between gastropod and protobranch hemocyanin than to cephalopod hemocyanin. Assuming a molecular clock, the last common ancestor of protobranch and gastropods lived 494 million +/- 50 million years ago, in conformity with fossil records from the late Cambrian.
Bergmann
Sandra
S
Institute of Zoology, Johannes Gutenberg University, 55099 Mainz, Germany.
Markl
Jürgen
J
Lieb
Bernhard
B
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D049872:Bivalvia / Q000235:genetics*; D018076:DNA, Complementary / Q000235:genetics*; D005786:Gene Expression Regulation; D006433:Hemocyanins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010802:Phylogeny; D011487:Protein Conformation
D018076:DNA, Complementary; D006433:Hemocyanins