Juan A. Subirana

Nuclear proteins from a somatic and a germinal tissue of the echinoderm Holothuria tubulosa

Abstract Histone fractions have been obtained from the haemal system and male gonads of the echinoderm Holothuria tubulosa . Fractionation was achieved by the methods of Johns [5], although some anomalies were found. The fractions were purified by precipitation in adequate solvent systems. The histones obtained are similar to those found in calf thymus, although f 1, and to a lesser extent f 2 b , show significant differences. In the gonads an additional minor component with high electrophoretic mobility is found. The other histone fractions appear to be identical with those obtained from somatic tissues. The relevance of this result towards an understanding of the role played by nuclear proteins in the sperm is discussed. Some of the acidic proteins present in the nuclei of these tissues are also preliminarily characterized.

Structural organization of sperm chromatin from the fish Carassius auratus

Abstract The structural organization of the histone-containing spermatozoa of the goldfish C. auratus is studied. Chemical fractionation shows that the five standard histones are present. Histone H1 subfractions are also similar to those found in other organisms. This is the first organism studied in which no peculiar basic protein is found in spermatozoa, since in all the organisms described up to now, unique protein components are always present, either alone or accompanied by somatic-like histones. Ultrastructural studies show that this chromatin is organized as a bundle of fibers of about 25 nm diameter, which upon spreading give the typical ‘beads-on-a-string’ appearance. Nuclease digestion demonstrates a repeat length of 205 base pairs (bp), slightly longer than in somatic tissues, but shorter than in echinoderm spermatozoa.

Nuclear proteins and the organization of chromatin in spermatozoa of Mythus edulis

The nuclei of sperm of the mussel Mytilus edulis contain three specific proteins: (1) a major protamine-like component (φ1); (2) a lysine-rich protein (φ3); and (3) a histone-like component (φ2b). The latter is characterized here for the first time. It is also shown that the φ3 protein does not contain modified lysines, but has some phosphorylated serines. Nuclease digestion of whole nuclei gives a continuous distribution of sizes for DNA, without any evidence of a nucleosome structure. X-ray diffraction indicates that DNA is organized as bundles of parallel molecules without any signs of either crystallinity or higher order structure.

Protamines and related proteins from spermatozoa of molluscs. Characterization and molecular weight determination by gel electrophoresis

The spermatozoa of most species of molluscs contain a mixture of proteins with compositions related to those of histones and protamines. The various components present in the spermatozoa of Cryptochiton stellerii, Donax trunculus, Mactra corallina and Gibbula divaricata have been isolated and characterized. The results obtained, taken together with those previously found (Subirana, J.A., Cozcolluela, C., Palau, J. and Unzeta, M. (1973) biochim. Biophys. Acta 317, 364--379), show that in all the molluscs studied one or more arginine-rich components are present. The molecular weight of these proteins varies vary much in different species and is usually much greater than in conventional fish protamines. Conventional histones, as well as lysine-rich proteins of low molecular weight, have also been found in ripe spermatozoa of several species. The molecular weights have been estimated by gel electrophoresis, using polymerized iridine as a standard.

Characterization of protamines from four avian species

No data are available on the protamines of birds, with the exception of galline. We have characterized the protamines from four species of birds belonging to four different orders. All of them have very similar properties. They have been purified by carboxymethylcellulose chromatography and analyzed with respect to amino acid composition and electrophoretic behaviour. They are very arginine‐rich proteins (63.4–67.3%) but do not contain lysine. Serine (12.0–18.2%), tyrosine (5.8–9.0%) and glycine (4.5–7.1%), along with arginine, make up the bulk of the amino acid residues in these molecules. The electrophoretic mobility of bird protamines in acetic acid‐urea‐polyacrylamide gels is intermediate between that of somatic histones and salmine. The molecular size, estimated from amino acid analysis and electrophoretic migration, is 65 ± 5 amino acid residues.

Morphology and crystalline structure of nylon-2/6

Abstract We have studied by X-ray diffraction and electron microscopy the crystalline structure of the copolyamide nylon-2/6 or poly(glycyl-e-aminocaproic acid). We have found a hexagonal form of the type known for polyglycine II with helical chains linked intermolecularly at an interchain distance of 4.79 A. We have obtained single crystals of this form from a formic acid solution. We have previously described this form in other nylons, so that it appears to be a new general structure for nylons, found in copolyamides which contain glycine or related monomers. We have also observed another modification made of hydrogen-bonded sheets which appear as ribbonlike lamellar crystals. The intersheet spacing measured for the latter form is 3.65 A, intermediate between those found in polyglycine I and in the α form of nylon-6.

Heterogeneity of the histone-containing chromatin in sea cucumber spermatozoa: Distribution of the basic protein Φ0 and absence of non-histone proteins

Nuclei of spermatozoa of the sea cucumber Holothuria tubulosa contain the five somatic-type histones plus a sperm-specific histone H1 and a unique basic protein phi 0, which is related to H1 in amino acid composition. No proteins of the High Mobility Group (HMG) type have been detected. The structure of this chromatin has been probed nuclease digestion. Its behaviour is anomalous, since two distinct fractions of chromatin are recovered from these spermatozoa, which differ either in the presence or absence of the sperm-specific proteins H1 and phi 0. This heterogeneous distribution is not found in conventional materials, such as calf thymus or chicken erythrocytes. Proteins H1 and phi 0 are not uniformly distributed and may be localized in special regions of chromatin. Fragments containing long stretches of nucleosomes lacking both proteins can be recovered. At the same time, the chromatin fractions which contain these two proteins are shown to be less soluble. When an extensive digestion of chromatin is carried out yielding only nucleosomes and small oligomers, the H1 and phi 0 proteins redistribute themselves on chromatin, the two proteins acting in a cooperative fashion in this process. Cross-linking experiments carried out in whole cells indicate a proximity of phi 0 and H1, whereas no crosslinks have been detected between phi 0 and any of the four nucleosomal histones. The phi 0 protein may thus play a role similar to histone H1 and be only loosely associated with nucleosomal histones, but contribute to the structuration of chromatin during spermiogenesis.

Comparison of protamines from freshwater and marine bivalve molluscs: Evolutionary implications

We characterized for the first time a protamine from the spermatozoa of a freshwater bivalve, Dreissena polymorpha. We found that it contains a protamine very similar in composition to those found in marine bivalves. We concluded that freshwater does not impose any restriction on the composition of sperm basic proteins. We compared our results with those recently published by other authors and proposed an evolutionary pathway for bivalve sperm proteins as originating from histone H1.

Presence of H2b histone in spermatozoa from marine gastropoda.

Abstract It is shown that spermatozoa from the marine snails Gibbula divaricata and Haliotis tuberculata contain a protamine plus a histone component. The composition of the latter is similar to H2 b histone from calf thymus.

Incorporation of glycine residues in even-even polyamides. Part II : Nylons 6,10 and 12,10

Sequential copolymers of glycine and nylons 6,10 and 12,10 have been prepared by incorporating single glycine units at both ends of the diamine. These polymers have been obtained by interfacial polymerization and subsequently characterized by infrared and n.m.r. spectroscopies together with thermal analysis. The structure and morphology of lamellar crystals have also been investigated by X-ray diffraction and transmission electron microscopy, both imaging and diffraction. Additional data have been obtained from uniaxially oriented fibres. Results described in this paper complete previous studies on the incorporation of glycine units in nylons derived either from ω-amino acids or from diamines and diacids.