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Biomedical subjects

G Dirheimer

Publications and source records attributed to G Dirheimer.

At least 163 records · Page 9Linked to original sources
PubMed1978

A calorimetric investigation of melting of tRNAAsp from brewer's yeast.

The thermodynamics of tRNAAsp unfolding was studied using a precision scanning microcalorimeter. The overall heat of melting was found to be about 55 J/g irrespective of the ionic strength and magnesium activity. The analysis of complex melting curves obtained in the absence of Mg2+ reveals four successive two-state transitions. The first was identified as the cooperative melting of the tertiary structure and the D region and the others as the melting of individual helical arms.

Aspartic Acid↗
PubMed1978

[Nucleotide sequence determination of yeast mitochondrial phenylalanine-tRNA].

The primary structure of mitochondrial tRNAPhe from Saccharomyces cerevisiae, purified by two-dimensional polyacrylamide gel electrophoresis, was determined using, standard procedures on in vivo 32P-labeled tRNA, as well as the new 5'-end postlabeling techniques. We propose a cloverleaf model which allows for tertiary interaction between cytosine in position 46 and guanine in position 15 and maximizes base pairing in the psi C stem, thus excluding the uracile in position 50 from base pairing in the psi C stem. Comparison of the primary structure of this tRNA with all other known procaryotic, chloroplastic or cytoplasmic tRNAsPhe sequences does not lead to any conclusion about the endosymbiotic theory of mitochondria evolution.

Base Sequence↗
PubMed1978

Neurospora crassa mitochondrial transfer RNAs.

Total mitochondrial tRNA from Neurospora crassa was characterized by base composition analysis, one- and two-dimensional gel electrophoreses and reversed-phase chromatography on RPC5. The guanosine + cytidine content was about 43%, as compared to 60% for cytoplasmic tRNA. The modified nucleoside content was low and about the same as that of total yeast mitochondrial tRNA, though the G + C content is very different. We found psi, T, hU, t6A, m1G, M2G, m22G. Neither the eukaryotic "Y" base, nor the prokaryotic s4U were present. On two-dimensional polyacrylamide gel electropherograms about 25 species were separated. One species for phenylalanine, two for leucine and two for methionine could be located. Neurospora crassa mitochondrial tRNA does not hybridize with yeast mitochondrial DNA.

DNA, Mitochondrial↗
PubMed1978

The primary structure of rabbit, calf and bovine liver tRNAPhe.

Highly purified tRNAPhe from rabbit liver, calf liver and bovine liver were completely digested with pancreatic ribonuclease and ribonuclease T1. The oligonucleotides were separated and identified. The tRNAPhe from rabbit liver and calf liver were partially cleaved with ribonuclease T1 or by action of lead acetate. We describe the analyses of the large fragments and the derivation of the primary structure of these mammalian tRNAsPhe.

Animals↗
PubMed1978

Primary structure of yeast mitochondrial DNA-coded phenylalanine-tRNA.

Mitochondrial tRNAPhe from Saccharomyces cerevisiae isolated by two-dimensional gel electrophoresis was sequenced by fingerprinting uniformly labeled 32 P-tRNA as well as by 5'-end postlabeling techniques. Its sequence was found to be: pG-C-U-U-U-U-A-U-A-G-C-U-U-A-G-D-G-G-D-A-A-A-G-C-m22G-A-U-A-A-A-phi-U-G-A-A-m1G-A-phi-U-U-A-U-U-U-A-C-A-U-G-U-A-G-U-phi-C-G-A-U-U-C-U-C-A-U-U-A-A-G-G-G-C-A-C-C-A. The secondary structure we propose, in order to maximize base pairing in the phiC stem and to allow tertiary interaction between G15 and C46, excludes U50 from base pairing giving a bulge in the phiC stem. No conclusion can be drawn concerning the endosymbiotic theory of mitochondria evolution by comparing the primary structure of mt. tRNAPhe with other sequenced tRNAsPhe. This mt.tRNAPhe lacks some of the structural elements reported to be involved in the yeast cytoplasmic phenylalanyl-tRNA ligase recognition site and cannot be aminoacylated by purified yeast cytoplasmic phenylalanyl-tRNA ligase.

Base Sequence↗
PubMed1978

Recognition of individual procaryotic and eucaryotic transfer-ribonucleic acids by B subtilis adenine-1-methyltransferase specific for the dihydrouridine loop.

Bulk tRNA from yeast and Rat liver can be methylated in vitro with -adenosylmethionine and B, subtilis extracts. The sole product formed is 1-methyladenosine (m1A). This tRNA (adenine-1) methyltransferase converts quantitatively the 3'-terminal adenosine-residue in the dihydrouridine-loop of tRNAThr and tRNATyr from yeast into m1A. Out of 16 eucaryotic tRNAs with known sequences 6 accepted methyl groups, all at a molar ratio of 1. These tRNAs have in common an unpaired adenosine-residue at the specific site in the sequence Py-A-A+-G-G-C-m2G. Out of 12 tRNAs from E. coli 6 served as specific substrates. These E. coli tRNAs also have an unpaired adenosine-residue at the 3'-end of the D-loop. Besides restrictions in primary structure intact secondary and tertiary structure is important for recognition of the specific tRNAs by the enzyme.

Bacillus subtilis↗
PubMed1977

[Primary structure of tRNA Thr 1a and b from brewer's yeast].

One of the two major species of brewer's yeast tRNA threonine (tRNA Thr 1) has been purified by countercurrent distribution followed by two chromatographic steps (respectively on a Sepharose 4B and a BD-cellulose column). Complete digestion with pancreatic and T1 RNases and a partial hydrolysis with T1 RNase followed by the isolation and determination of the nucleotide sequences of the resulting fragments permitted the derivation of its primary structure. tRNA Thr 1 is in fact a mixture of two subspecies differing only by a A49-U65 base pair in 50 per cent of the molecules which is replaced by a G49-C65 pair in the other 50 per cent. These two subspecies consist of 76 nucleotide residues including 14 minor nucleotides. They show a characteristic m3C at the 3'terminal end of the anticodon loop, an anticodon I-G-U followed by t6A and C48, uncompletely modified (50 per cent) to m5C within the 5 nucleotides long extra-arm. The minor nucleotides m2G m2 2G are located at positions in which they generally occur in the tRNA structures as does m1A within the T-psi-C loop.

Base Sequence↗
PubMed1977

Aminoacylation of tRNA Trp from beef liver, yeast and E. coli by beef pancrease tryptophan-tRNA ligase. Stoichiometry of tRNATrp binding.

The Michaelis constants and the maximum velocities in the aminoacylation reaction of tRNATrp from beef liver, yeast and E. coli by pure beef pancreas tryptophan-tRNA ligase show that this mammalian enzyme recognizes and charges the two eucaryotic tRNAs with the same efficiency. The rate of aminoacylation of the procaryotic tRNATrp by the enzyme is three orders of magnitude lower. The pH optimum of aminoacylation is 8 for both eucaryotic tRNAs. The optimum magnesium concentration is different. The rate is maximum when magnesium concentration is stoichiometric to ATP concentration for tRNATrp from beef liver and 10 mM above ATP concentration for tRNATrp from yeast. The number of binding sites on the enzyme for the two eucaryotic tRNAs has been measured by equilibrium filtration on Sephadex G-100 and found equal to two.

Adenosine Triphosphate↗
PubMed1977

Study of yeast mitochondrial tRNAs by two-dimensional polyacrylamide gel electrophoresis: characterization of isoaccepting species and search for imported cytoplasmic tRNAs.

By two-dimensional polyacrylamide gel electrophoresis, yeast mitochondrial tRNA is fractionated into 27 major species. All but 6 of them migrate distinctly from cytoplasmic tRNAs. Migration of mitochondrial DNA-coded mitochondrial tRNAs shows the occurence of only one cytoplasmic tRNA in mitochondria. Several mitochondrial tRNA spots are identified on the electrophoregrams, some of them show isoaccepting species (Val, Ser, Met, Leu). It is suggested that there are sufficient mitochondrial tRNA genes on yeast mitochondrial DNA to allow mitochondrial protein biosynthesis by the mitochondrial tRNAs alone. Guanosine + Cytidine content and rate base composition are reported for some individual species. Mitochondrial tRNAPhe lacks Ribothymidine.

Electrophoresis, Polyacrylamide Gel↗
PubMed1977

Methylation of an adenosine in the D-loop of specific transfer RNAs from yeast by a procaryotic tRNA (adenine-1) methyltransferase.

tRNA (adenine-1) methyltransferase occurs in Bacillus subtilis. Eucaryotic tRNAThr and tRNATyr from yeast in which 1-methyladenosine (m1A) is already present in the TpsiC loop, can be methylated in vitro with S-adenosylmethionine and B. subtilis extracts. Each of the specific tRNAs accepts 1 mol of methyl groups per mol tRNA. The enzyme transforms into m1A the 3'-terminal adenylic acid residue of the dihydrouridine loop, a new position for a modified adenosine residue in tRNA. Both tRNAs have the sequence Py-A-A-G-G-C-m2(2)G in the D-loop and D-stem region. Other tRNAs with the same sequence in this region also serve as substrates for the tRNA (adenine-1) methyltransferase.

Adenine Nucleotides↗