Amino-acid sequence of Tetrahymena histone H4 differs from that of higher eukaryotes.

Abstract

A partial amino-acid sequence of Tetrahymena histone H4 has been determined and differs significantly from the sequence of calf or pea H4. The amino terminus of Tetrahymena H4, unlike that of other H4s so far examined, is not acetylated. Of 66 residues determined, one is a single-residue insertion, one a single-residue deletion, and thirteen are amino-acid replacements with respect to the calf thymus H4 sequence. Most Histone H4 is the most highly conserved protein known. The complete sequences of H4 of calf, pea, pig, and rat (1-4) are identical except for replacements in the pea sequence of isoleucine for valine and arginine for lysine at positions 60 and 77, respectively. Partial sequencing of H4 from other sources (5-7) has revealed only one additional amino-acid replacement: cysteine for threonine at residue 73 of the sea urchin sequence (7). Whether this extreme conservatism extends to H4 of all eukaryotes remains to be seen, but electrophoretic and compositional analyses of H4s from a wide variety of species (8-13) suggest that the sequence of H4 has remained essentially constant during the evolution of most eukaryotic organisms.Histone H4 of Tetrahymena thermophila shares many biochemical features with H4 of higher eukaryotes. The electrophoretic behavior of Tetrahymena H4 both on sodium dodecyl sulfate (NaDodSO4) and on acetic acid/urea polyacrylamide gels is indistinguishable from that of calf thymus H4, and both histones behave similarly in purification procedures involving either exclusion chromatography or differential solubility (14). Several minor differences have been reported, however, in the amino-acid compositions of Tetrahymena and calf H4 (14).We have recently observed that Tetrahymena H4 fails to substitute efficiently for calf H4 in nucleosome reconstitution experiments (15) and complexes less strongly than calf H4 with both calf and Tetrahymena H2B and H3 (16). These observations have led us to investigate the primary structure of this histone. We report here that substantial differences exist between Tetrahymena H4 and H4 of higher eukaryotes. MATERIALS AND METHODSHistone H4 was obtained from T. thermophila, strain B VII. Nuclei were isolated and whole histone was extracted as described (17). Histone H4 was purified by chromatography on Bio-Gel P-60 followed by chromatography on carboxymethyl-cellulose (14). Calf thymus H4 was purified by similar procedures. Purified fractions were dialyzed exhaustively against distilled, deionized H20 and lyophilized. Both proteins were greater than 98% pure as estimated by NaDodSO4/ polyacrylamide gel electrophoresis (18,19).Cleavage at methionine residues was obtained by exposure to 10-to 300-fold molar excess of cyanogen bromide (CNBr) in 70% formic acid for 24 hr at 23°C in the dark. CNBr peptides were analyzed on NaDodSO4/polyacrylamide gels by the method of Laemmli (18) modified as follows: the running gel was 20% acrylamide/1% bisacrylamide, and both running and stacking gels contained 8 M urea.Automated Edman degradations were perform...

Unusual pattern of ribonucleic acid components in the ribosome of Crithidia fasciculata, a trypanosomatid protozoan.

“…To date, however, relatively few eucaryotic ribosomal systems have been studied in detaiL and these do not represent a phylogenetically diverse sampling. Since data from protein sequence analysis (8,13) suggest an ancient evolutionary separation, substantially antedating the divergence of plants and animals (31), between the protozoa and higher eucaryotes, one might anticipate that some particularly distinctive features of ribosome structure or function (or both) will emerge from studies of these (and perhaps other) members of the Kingdom Protista. This expectation is borne out by the results presented in this and a previous (14) publication, which demonstrate that in its structural properties, the ribosome of C. fasciculata is the most "atypical" eucaryotic ribosome yet described.…”

A single histone acetyltransferase from Tetrahymena macronuclei catalyzes deposition-related acetylation of free histones and transcription-related acetylation of nucleosomal histones.

“…To analyze the sites on a free histone substrate, un-acetylated macronuclear H4 was purified from long acidurea gels (see Materials and Methods), acetylated in vitro under standard conditions, and reanalyzed on a second long acid-urea gel. Reaction products corresponding to mono-or diacetylated H4 were excised, eluted, and subjected to automated microsequencing (note that Tetrahymena H4 does not have a blocked amino terminus as do most other H4s; see Glover and Gorovsky, 1979). It is clear that the lysine at residue 11 is the exclusive site of acetylation in monoacetylated H4 (Fig.…”

Characterization of Histone H3/H4 Gene Region and Phylogenetic Affinity of Ichthyophthirius multifiliis Based on H4 DNA Sequence Variation

“…Yet, among 29 species of Tetrahymena and Glaucoma chattoni, Sadler and Brunk (1992) detected no amino acid replacements within the amino-terminal third of the H3 coding region, further suggesting that selective pressures acting on ciliates are different from those acting on other eukaryotes. Together, these data suggest that the tempo and mode of evolution of eukaryotic histone genes in ciliated protozoans are different from those in higher eukaryotes (Glover and Gorovsky, 1979;Bannon et al, 1984;Horwitz et al, 1987;Brunk et al, 1990;Sadler and Brunk, 1992;Bernhard and Schlegel, 1998).…”