Observations on the nature and the interaction of a carotenoprotein and a cytochrome from the eggs of a prosobranch mollusc
Evidence is given for interaction of the carotenoglycoprotein ovorubin and a haemoprotein, spectroscopically indistinguishable from the cytochrome-like helicorubin of Helix pomatia found in the egg jelly of the prosobranch Pomacea canaliculata. Ovorubin catalyses reduction of helicorubin by NADH', NADPH, GSH and ascorbate. Addition of ATP, ADP and AMP to the egghomogenate elicits changes in the Fe/Fe ratio of the haemoprotein. These are dependent on the initial ratio and the nucleotide concentration. ATP synthesis could sometimes be shown in homogenates of eggs developed for 100 hrs and correlated with redox changes in helicorubin. No ATP synthesis is observed with ovorubin-free homogenates or those of freshly laid eggs. Returning ovorubin to homogenates freed from this protein does not restore ATP syrrthesis. Egg homogenates dephosphorylate ATP and ADP. Some 10% of this activity remains in chromatographically prepared ovorubin. Extensive purification results in loss of this activity. Ovorubin, but not its apoprotein, undergoes spectral changes, reversed by light, on exposure to carbon monoxide. E.S.R. spectra of ovorubin show signals at g = 1.96 and 4.2, suggestive of non-haem iron. Electron micrographs of ovorubin, together with a sub-unit structure determined by SDS-PAGE, are used to suggest a molecular model. Helicorubin from P. canaliculata has been purified and found to be a basic protein. Ovorubin, added to oxidised helicorubin or cytochrome c from horse heart or Candida, gives partial reduction in dark. In visible light, reduction is virtually complete. With apoprotein partial reduction occurs in light or dark to the degree found with the carotenoprotein in the dark. Reduction of cytochrome by ovorubin or its apoprotein is competitively inhibited by superoxide dismutase indicating generation of Anaerobically in the dark, reduction of cytochrome by ovorubin and apoprotein reaches about one third the level with ovorubin aerobically in the light and one half that with ovorubin aerobically in the dark. These observations suggest the presence of three redox centres on ovorubin. The functional significance of these interactions, especially during embryonic development, is discussed.