Primary Structure of Histone H2A from Gonad of the Sea Urchin Psammechinus miliaris

Abstract

The complete amino acid sequence (325 residues) of sea urchin histone H2A has been established by structural studies of peptides derived from tryptic and chymotryptic cleavage of the maleylated protein and from thermolysin cleavage of the intact protein. By comparison with calf homologous histone, the basic amino-terminal and carboxy-terminal parts of the protein show 11 substitutions and 4 deletions. The remainder of the sequence, mostly hydrophobic, is almost completely unchanged.Among the five histones characterized in the chromatin of the eukaryotes, the histones H3 and H4 show a remarkable structural stability throughout the evolution [1,2] whereas the other histones present a structural variability, particularly accentuated in the histone H1.This variability first evidenced by comparative electrophoretic studies of whole histone from vertebrates [3] has been confirmed by sequence determinations [1,4-121. Recently, the primary structures of histone H2B1 and H2B2 from sperm of the sea urchin Parechinus angulosus have been reported [13,14]. The authors have shown that considerable variation occurred in the amino-terminal part of these proteins and that the hydrophobic carboxy-terminal region was almost completely invariable.In this paper, we report the primary structure of histone H2A from gonads of sea urchin Psammechinus miliaris. This amino acid sequence has been established from the data provided by tryptic and chymotryptic peptides of maleylated protein and by thermolysin peptides of intact protein. MATERIALS AND METHODS MaterialsTrypsin [treated with ~-(l-tosylamido-2-phenyl)-ethyl chloromethyl ketone] and chymotrypsin (premium quality WCDS) were purchased from Worthington. Carboxypeptidases A and B (treated with diisopropylphosphofluoridate) and thermolysin were purchased from Merck. Aminopeptidase M and carboxypeptidase C were obtained from Rohms and Haas.Phenylisothiocyanate, propan-1 -01 and dimethylallylamine were Sequanal grade from Pierce. Benzene and ethyl acetate for sequential analysis were purchased from S.D.S. (Peypin, France). Ethylmercaptan and propionic acid were obtained from Fluka. All other reagents were of the purest grade commercially available.

Complete Amino‐Acid Sequences of DNA‐Binding Proteins HU‐1 and HU‐2 from Escherichia coli

“…The last fractions eluted from the Sephadex were indeed a complex mixture of small peptides which were further fractionated by ion-exchange chromatography on Chromobeads P column (Technicon Corporation) as described in [12]. The tryptic and chymotryptic peptides were directly fractionated by ionexchange chromatography.…”

The Primary Structure of Histone H2A from the Sperm Cell of the Sea Urchin Parechinus angulosus

“…This histone, as other H2A histones [3,21] has no free amino terminus. However using an N -+ 0 acyl shift followed by coupling of phenylisothiocyanate to the newly generated amino groups, the methyl ester protein was sequenced for 32 steps (Table 3- 1) and the N-terminal peptide TH-11-5 (Table 3-2) for 11 steps.…”

“…Histones H2A also show an intermediate degree of variability which occurs mostly in the amino-terminal and carboxyl-terminal regions while the hydrophobic central region is very conservative [I]. The primary structure of histone H2A from gonads of the sea urchin Psammechinus miliuris has been published [ 3 ] . Since the amino-terminal regions of histone H2B from this species of sea urchin [4] differ appreciably from the same regions of histone H2B from Parechinus anguIosus [5 -71, the sequence of histone H2A from sperm of P. angulosus has been completed and compared to that of Ps.…”

“…by s u b t r a c t i o n * 1 mq h l s t o n e H4 was added w l t h the p e p t l d e and the p e p t l d e program Was used used to determine the sequence are given in Tables 3-1 to [3][4][5][6][7][8][9][10][11][12][13][14][15][16][17]. This histone, as other H2A histones [3,21] has no free amino terminus.…”

“…The peptides adjoining at these residues have been aligned by analogy to the sequence of the H2A histone of the sea urchin Psumrneclzinus miliuris [3].…”