No abstract

Analysis of the total protein of the mature sperm of the bivalve mollusc Ensis minor (razor shell) using gel electrophoresis, amino acid analysis, nuclear magnetic resonance, circular dichroism and trypsin digestion, show it to contain all five histones plus a protamine-likq protein. The histones H3, H 4 and probably H2A are similar to those from calf thymus or sea urchin sperm, but $he putative H2B appears to have a very high molecular mass (about 20 kDa). The histone H 1 molecule is unusual, having little or no proline and 8-10 residues of histidine. The protamine-like species is rich in both lysine as well as arginine and is of much higher molecular mass than fish sperm protamines. Nucleosomes containing the four core histones have been prepared and the nucleosomal repeat shown to be 200 f 5 base pairs. Checks for the absence of contaminating cells reinforce the conclusion that a histone-containing nucleosomal structure co-exists with a protamine-like protein in this sperm chromatin.Electrophoretic and amino acid characterisation of the proteins from the sperm of several species of mollusca has been published [I -61. All authors, with the exception of Balhorn et al. [2], have concluded that both protamine-like proteins and histones are present in the protein mixtures extracted from such sperm cells. In order to clarify the nature of these proteins further and to understand the apparent co-existence of protamine-like proteins and histones, we decided to concentrate out attention on a single species of bivalve mollusc Ensis minor (razor shell) since the evidence for the presence of both histones and protamines is strong for this class [l-61. Both physical and biochemical characterisation of proteins extracted from ripe sperm has been carried out. In addition to a protamine-like species a full range of histone types have been found, including an unusual H I-type histone. Apparently normal nucleosomal core particles have been extracted. MATERIALS AND METHODS Sperm preparation and protein extractionSpecimens of Ensis minor were collected from the Adriatic Sea between November and March, the period of maximum sperm production. The majority of the protein characterisation is from a March collection. The gonadal tissue was opened with a blade and mature sperm shaken gently into filtered sea water at 0 -4 "C made 0.5 mM in phenylmethylsulphonyl fluoride. The suspension was filtered first through three layers of surgical gauze and then through a plankton net. Sperm were recovered by low-speed centrifugation. Microscopic examination indicated that 95 % of the material was fast-moving mature sperm. After washing twice in 4.25% KCl, basic Abbreviations. SDS, sodium dodecyl sulphate; CD, circular dichroism.proteins were directly extracted with 0.3 M H,S04 using a blender for three 2-min periods. In some preparations 0.75 % Triton X-100 in 0.25 M sucrose, 50 mM Tris/HCl, pH 7.5, 10 mM MgCl,, 0.1 mM phenylmethylsulphonyl fluoride, was used to wash sperm so as to remove tails. Sperm heads were then washed twice with the same ...

No abstract

No abstract

No abstract