The Structure of Sea‐Urchin‐Sperm Histone φ1 (H1) in Chromatin and in Free Solution

Abstract

The lysine-rich H1-type histone 41 from the sperm of the sea urchin Arbaciu lixula has been subjected to tryptic digestion in free solution at high ionic strength. Tw trypsin-resistant peptides G41 and LG41 have been isolated, comprising 81 and 93 amino acids respectively, i.e. about a third of the intact histone. Circular dichroism and 270-MHz proton NMR have been used to demonstrate that the smaller peptide G41 contains all the secondary and tertiary structure of intact histone 41. It is concluded that the resistant peptides represent a compact folded portion of the histone 41 chain, whilst the remaining two-thirds is disordered in free solution. Trypsin digestion of Arbacia lixulu nuclei, under conditions where histone 41 remains tightly bound to the chromatin, leads to the same resistant peptide G41. From this it is concluded that in chromatin the chain region corresponding to G41 is likewise folded and inaccessible to trypsin, whilst the remaining exposed residues are located periferally and probably in an extended conformation. The rather unusual H1-type histone 41 from sea urchin sperm thus has a three-domain structure like calf thymus H1 and chicken erythrocyte H5. The present data emphasise the fact that this threedomain structure exists in chromatin and is not merely a free-solution artefact.Considerable progress in the understanding of chromatin structure has been made in recent years through the use of nucleases to probe DNA conformation. The use of proteases to study the conformation of chromosomal proteins has received less attention. Experiments on the trypsin digestion of the core histones in chromatin [1,2] have led to the conclusion that the N-terminal regions of these histones are preferentially digested. Although the structures underlying the high exposure of these parts of the core histones are not known, these results are in accord with the concept of a structural distinction between the N-terminal and C-terminal regions of the histones that has followed from studies of the (H3/H4)2 tetramer and (H2A/H2B)2 dimer in free solution [3 -51. In that work it was shown that the N-terminal regions of the histone complexes are not compacted, but free and mobile in solution. A similar approach has been taken in the study of the conformation of histone H1 in free solution. Tryptic digestion was shown to result in preferential digestion of 35 N-terminal residues and all the C-terminal half of the molecule beyond residue 120 [6]. The remaining central portion of the molecule was shown to contain all the secondary and tertiary structure of intact H1 and moreover to retain this structure after removal of the N-terminal and C-terminal regions. A three-domain structure containing a single central folded region has also been demonstrated for histone H5 in a similar manner [7], although in this case the N-terminal region was 21 residues long, rather than 35 residues.Free solution studies of histone conformation are subject to the uncertainty that since the histone has been removed from the other components wi...

DNA and histone H1 interact with different domains of HMG 1 and 2 proteins.

“…We used two proteases to test in vitro a functional correlation with the asymmetry observed in the sequence. Trypsin has provided information about the structure of chromatin proteins, both core histones (Weintraub, 1975;Whitlock and Simpson, 1977) and histone HI (Hartman et al, 1977) and related proteins (Aviles et al, 1978;Puigdomenech et al, 1980). This enzyme has been used to test the domain hypothesis for HMG-1, -2 and -E (Paulau et al, 1980;Reeck et al, 1982;Cary et al, 1983).…”

“…We have examined the domain structure for HMG-1 and -2 from calf thymus following a strategy similar to that used to show the domain structure of histone HI (Hartman et al, 1977;Aviles et al, 1978;Puigdomenech et al, 1980). The proteins were digested with proteases and the products of the digestion analyzed.…”

Preparation and characterization of histone H1 from the sperm of the sea-urchin Sphaerechinus granularis

“…The physical characterization described above indicates that both the secondary structure and tertiary structure of S. granularis histone H1 are very similar to those of A. lixula histone HI, despite significant compositional differences. The folding domain of H1 histones in which the secondary and tertiary structures are located is of approximately 80 residues (Hartman et al, 1977;Puigdomenech et al, 1980;Allan et al, 1980) and is centrally located. Preliminary digestion studies indicate that S. granularis histone H 1 also contains a trypsinresistant domain of molecular weight approx.…”

Precise elimination of the N-terminal domain of histone H1

“…86 residues in calf thymus HI) and a long unstructured C-terminal domain of very basic composition comprising almost half the molecule (approx. 93 residues in calf thymus HI) (Chapman et al, 1976;Hartman et al, 1977;Puigdomenech et al, 1980;Barbero et al, 1980;Aviles et al, 1978). By reconstituting several HI peptides on to chromatin depleted of its original Hi, it was shown that the central globular domain alone is able to close two full turns of nucleosomal DNA.…”