Observations on the structure and function of the haemoglobin from the cuticle ofNippostrongylus brasiliensis(Nematoda)

Abstract

SUMMARYThe fluid layer within the cuticle of the nematodeNippostrongylus brasiliensiscontains haemoglobin. The absorbance spectra of the haemoglobins extracted from whole nematodes were measured spectrophoto-metrically and the spectra of cuticular haemoglobin in individual living nematodes was obtained by microdensitometry. Adult female nematodes were confined under cover-slips in oxygenated saline and measurements of optical density of a small area of the cuticular haemoglobin were taken at 10 sec intervals. The optical density decreased over a period of 4 min at 543 nm and increased at 553 nm indicating deoxygenation of the cuticular haemoglobin; irrigation with fresh saline restored the haemoglobin to its oxygenated state. This demonstrates that cuticular haemoglobin loads and unloads oxygen in the living animal. Isoelectric focusing showed that mature nematodes have haemoglobins which are isoelectric around pH 7·0 and at pH 9·8. Isolated cuticular fluid contained the haemoglobin which was isoelectric at pH 9·8, but no other haemoglobin, and immature adults, which do not have cuticular haemoglobin, contained only those haemoglobins which were isoelectric at pH 7·0. Sodium dodecyl sulphate polyacrylamide gel electrophoresis and column chromatography showed that the molecular weight of the cuticular haemoglobin sub-unit is 16·9–17·5 × 103Daltons and that the haemoglobin exists primarily as a tetrameric molecule with a molecular weight of 65–75 × 103Daltons. The presence of cuticular haemoglobin may allow adultNippostrongylusto exploit oxygen-deficient areas within the environment of the rat's intestine where it would otherwise be unable to survive for prolonged periods.