Haemocyanin mRNA from arthropod and mollusc origin. Evidence for a multi-unit structure in mollusc haemocyanin mRNA.

mRNA from two molluscs, the snail Levantina hierosolima and the cuttlefish Sepia officinalis, and one arthropod, the scorpion Leiurus quinquestriatus, were found to contain species that hybridize with an oligodeoxynucleotide sequence corresponding to the six-amino-acid sequence His-His-Trp-His-Trp-His postulated to carry binuclear copper in arthropod haemocyanins. The duplexes formed between the oligodeoxynucleotide and mRNA from Leiurus and Levantina had similar 'melting' temperatures, namely 46.5 and 44.5 degrees C respectively. The hybridizable mRNA species were identified with mRNA that codes for haemocyanin, Hc mRNA. Electrophoresis under denaturing conditions of Leiurus Hc mRNA gave a single band corresponding to 2.3 kb (kilobases), consistent with the value of 75 000 reported for the Mr of the polypeptide chain of arthropod haemocyanin. Electrophoresis of mollusc Hc mRNA yielded RNA bands corresponding to 9.4, 6.7, 4.0 and 1.7 kb for Levantina and 9.5, 2.8 and 1.7 kb for Sepia. The 9.4 and 9.5 kb species represent authentic Hc mRNA and are consistent with an Mr of 350 000 for molluscan haemocyanin polypeptide chain. The faster-moving RNA bands are attributed to Hc mRNA cleavage by nucleases during isolation of mRNA. Analysis of the electrophoretic band pattern indicates a multi-unit structure for mollusc Hc mRNA.

Immunological correspondence between arthropod hemocyanin subunits. I. Scorpion (Leiurus, Androctonus) and spider (Eurypelma, Cupiennius) hemocyanin.

The hemocyanins of the scorpions Leiurus quinquestriatus and Androctonus australis, the tarantula Eurypelma californicum (all 24-mers), and the lycosid spider Cupiennius salei (dodecamer) were dissociated into subunits, the subunits isolated and studied by two-dimensional immunoelectrophoresis for interspecific cross-reactivities. Androctonus hemocyanin yielded a pattern of 8 subunit types in agreement with data from Lamy et al. (1979, Arch. Biochem. Biophys. 193, 140-149). Leiurus hemocyanin is also composed of 8 immunologically distinct subunits which could be assigned to the pattern of Androctonus in a subunit-to-subunit correlation. The subunit designations 1 to 6 of Lamy et al. could be adopted for both scorpion hemocyanins; however, in the present communication, Lamy's subunits 3A/3B are designated as 3'/3", because we could not unequivocally decide if 3' = 3A and 3" = 3B or vice versa. The 7 subunit types a to g of Eurypelma hemocyanin could be correlated with the scorpion hemocyanin subunits as follows: a = 3', b = 5B, c = 3C, d = 5A, e = 6, f = 2, g = 4. Additional cross-reactivities were detected between e/4, and f/5A, respectively. No subunit of Eurypelma hemocyanin is homologous to scorpion 3", which could not be precipitated by anti-Eurypelma antiserum. Antiserum against Cupiennius hemocyanin precipitated subunit f of Eurypelma and subunits 2 and 5A of scorpion hemocyanin. The published models of quaternary structure and a possible subunit phylogeny of arachnidan hemocyanins are discussed in view of the present results.