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Biomedical subjects

H Busch

Publications and source records attributed to H Busch.

At least 379 records · Page 21Linked to original sources
PubMed1975

Nuclear ribonucleoprotein complexes containing U1 and U2 RNA.

Nuclear ribonucleoprotein (RNP) complexes that contain the U1 and U2 RNA of chromatin of Novikoff hepatoma cells were extracted with 0.01 M Tris-HCl (pH 8.0) after the nuclei were initially washed with 0.075 M NaCl and 0.025 M EDTA (pH 8.0). These RNP complexes were purified by chromatography on Sepharose 6B columns and centrifugation on sucrose density gradients. The identity of the U1 and U2 RNA in these particles was established by their electrophoretic mobility in polyacrylamide gels and their T1 RNase fingerprints which were identical with those of authentic U1 and U2 RNA (R. Reddy et al. (1974), J. Biol. Chem.249, 6486-6494; H. Shibata et al. (1974), Mol. Cell. Biochem. 4, 3-19). The nuclear riboncleoproteins had a buoyant density of 1.47 g/ml in CsCl gradients. Two-dimensional polyacrylamide gel electrophoresis of their proteins showed these RNP complexes contain 10 polypeptide spots, of which two are phosphorylated in vivo.

Animals↗
PubMed1975

Isolation and characterization of protein A24, a "histone-like" non-histone chromosomal protein.

In earlier studies, the nucleolar levels of protein A24 were found to be markedly decreased in the nucleolar hypertrophy induced by thioacetamide or during liver regeneration (Ballal, N.R., Goldknopf, I.L., Goldberg, D.A., and Busch, H. (1974) Life Sci. 14, 1835-1845; Ballal, N.R., Kang, Y.-J., Olson, M.O.-J., and Busch, H.J. Biol. Chem. 250, 5921-5925). To determine the role of protein A24, methods were developed for its isolation in highly purified form. Milligram quantities of highly purified protein A24 were isolated from the 0.4 N H2SO4-soluble proteins of calf thymus chromatin by exclusion chromatography on Sephadex G-100, followed by preparative polyacrylamide gel electrophoresis. Protein A24 was highly purified as shown by its migration as a single spot on two-dimensional polyacrylamide gel electrophoresis, its single NH2-terminal amino acid, methionine, and the production of approximately 50 peptides by tryptic digestion. Like histones 2A, 2B, 3, and 4. A24 was extractable from chromatin with 0.4 N H2SO4 or 3 M NaCl/7 M urea, but unlike most non-histone proteins or histone 1, protein A24 was not extracted with 0.35 M NaCl, 0.5 M HClO4, or 0.6 M NaCl. Protein A24 was present in only 1.9% of the total amount of histones 2A, 2B, 3 and 4; its molecular weight is 27,000.

Amino Acids↗
PubMed1975

Physical and chemical characterization of purified ovalbumin messenger RNA.

Preparative agarose gel electrophoresis under denaturing conditions has been successfully employed to purify large quantities of ovalbumin mRNA from hen oviducts. The mRNA thus prepared is physically homogeneous based on its migration as a single component on electrophoresis in both analytical acid-urea agarose gels and formamide-containing, neutral polyacrylaminde gels; it also sediments as a single peak in sucrose gradients containing 70% formamide. The mRNA is chemically free of ribosomal RNA contamination since its oligonucleotide fingerprint map after complete T1 ribonuclease digestion contains no detectable specific large oligonucleotide markers of ribosomal RNAs. It is also not contaminated by other biologically active messenger RNAs because, when it is added to the cell-free wheat germ translation system, the only protein product synthesized is ovalbumin as analyzed by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and specific immunoprecipitation. Ovalbumin mRNA has a nucleotide composition of 32.3% A, 21.0% G, 25.7% U, and 20.7% C [(A+U)/(G+C) equal 1.41]. The mRNA contains a heterogeneous poly(A) tract ranging from 20 to 140 residues with a number average chain length of 62 adenylate residues. The molecular weight of the sodium salt of the purified mRNA is approximately 650,000 +/- 63,000, corresponding to a chain length of 1890 +/- 180 nucleotides, as determined by electron microscopy under completely denaturing conditions. This value is in close agreement with the values obtained from: (a) sucrose gradient centrifugation in the presence of 70% formamide; (b) evaluation of poly(A) content in the mRNA and the number average chain length of its poly(A) tract; and (c) sedimentation velocity studies in the presence of 3% formaldehyde. When 125I-labeled ovalbumin mRNA is allowed to hybridize with a large excess of chick DNA, the observed kinetics of hybridization reveal no appreciable reaction between the mRNA and the repeated sequences of the chick DNA, although the mRNA appears to be approximately 600 nucleotides longer than necessary to code for ovalbumin. It thus appears that the entire ovalbumin mRNA is primarily transcribed from a unique sequence in the chick genome.

Adenine Nucleotides↗
PubMed1975

Changes in nucleolar proteins and their phosphorylation patterns during liver regeneration.

Two-dimensional polyacrylamide gel electrophoresis of nucleolar proteins of rat liver revealed marked changes at various times after partial hepatectomy. Some of the non-histone protein spots including A11, A24, A25, C13, and C14 decreased in size and density. Others, including A15, B13, B16, B18, B24-25, B27, B33, B34, C1, C2, C8, C11, C17, and C23-24 were markedly increased in size and density. In vitro labeling of the nucleoli using [gamma-32P]ATP and subsequent analysis of the proteins by two-dimensional gel electrophoresis and autoradiography indicated that the uptake of 32P into proteins increased greatly during regeneration. In addition, the relative labelling of various spots changed throughout the regeneration process. With spot B33 as a reference, there was increased labeling of spots A1P, B2, B5, B24-25, C2, and C23-24 at 8 hours after hepatectomy. On the other hand, the labeling of spots A8P, A16, A20, B9, and B13 decreased at 8 hours after hepatectomy. Since the primary role of the nucleolus is the snythesis of ribosomal precursors, these changes in protein content and phosphorylation are presumably primarily associated with the increased processing and transport of peribosomal ribonucleoproteins in regenerating liver.

Adenosine Triphosphate↗
PubMed1975

The primary nucleotide sequence of nuclear U-2 ribonucleic acid. The 5'-terminal portion of the molecule.

The nuclear U-2 RNA which is highly modified (Reddy, R., Ro Choi, T.S., Henning, D., Shibata, H., Choi, Y.C., and Busch H. (1972) J. Biol. Chem. 247, 7245-7250) contains 13 pseudouridylic acid residues, 10 2'-O-methylated nucleotides and two modified bases including N-2,2, 7-trimethyl guanylic acid in its 5'-terminal portion (69 nucleotides). With the determination of this sequence and its overlap with the 3' portion of the molecule (nucleotides 70 to 196), the over-all nucleotide sequence of this RNA is:(see article). The concentration of modified nucleotides in its 5' portion is greater than for any RNA sequenced thus far.

Base Sequence↗
PubMed1975

Nucleotide sequence of U-2 ribonucleic acid. The sequence of the 5'-terminal oligonucleotide.

The nucleotide sequences were determined for the 5'-oligonucleotides obtained by complete pancreatic RNase digestion (P25) and complete T1 RNase digestion (T27) of U-2 RNA. Complete digestion of oligonucleotide P25 with snake venom phosphodiesterase produced pm3 2,2,7G, pAm, pUm, and pCp in approximately equimolar ratios. Partial digestion of these oligonucleotides with snake venom phosphodiesterase produced -Um-C-Gp and pAm-Um, indicating the sequence of the 3'-terminal portion of the 5'-oligonucleotide is pAm-Um-C-Gp. The 5'-terminal oligonucleotide did not contain a 5'-phosphate and no free nucleoside was released from the 5' end by venom phosphodiesterase digestion. Since free pm3 2,2,7G was released by digestion with nucleotide pyrophosphatase and limited digestion with snake venom phosphodiesterase, this nucleotide is apparently linked to pAm in a pyrophosphate linkage. Mass spectrometry and thin layer chromatography in borate systems showed the ribose of m3 2, 2, 7G contains no 2'O-methyl residue. Moreover, the finding that the ribose of m3 2, 2, 7G was oxidized by NaIO4 and reduced by KB3H4 in intact U-2 RNA rules out other linkages involving the 2' and 3' positions. Accordingly, it is concluded that the structure of the 5'-terminal pentanucleotide of U-2 RNA is(see article).

Alkaline Phosphatase↗
PubMed1975

Maturation pathway for Novikoff ascites hepatoma 5.8 S ribosomal ribonucleic acid. Evidence for its presence in 32 S nuclear ribonucleic acid.

Evidence that 32 S nRNA contains 5.8 S rRNA was provided by studies on specific oligonucleotide sequences of these RNA species. Purified 32P-labeled 5.8 and 28 S rRNA and 32 S RNA were digested with T-1 ribonuclease, and the products were fractionated according to chain length by chromatography on DEAE-Sephadex A-25 at neutral pH. The oligonucleotides in Peak 8 were treated with alkaline phosphatase and the products were separated by two-dimensional electrophoresis on cellulose acetate at pH 3.5 and DEAE-paper in 7% formic acid. Seven unique oligonucleotide markers for 5.8 S rRNA including the methylated octanucleotide A-A-U-U-Gm-G-A-Gp were present in 32 S RNA but were not found in 28 S rRNA, indicating that 5.8 S rRNA is directly derived from the 32 S nucleolar precursor. These studies confirm a maturation pathway for rRNA species in which 32 S nucleolar RNA is a precursor of 5.8 S rRNA as well as 28 S rRNA.

Alkaline Phosphatase↗
PubMed1975

[Practice and further development of the AMP-system (a report about a symposion and a seminar) (author's transl)].

During two days in November 1974 TV-tapes of psychiatric interviews were demonstrated and the psychopathology was documented on AMP-record sheet 3. It was a purpose of the discussion to get stimulation for the improvement of the AMP-manual and to decrease the interrater-variance. It was only the first tape which was not followed by a discussion. This film was presented a second time at the end of the seminar in order to examine the training effects. Both record-sheets were compared with each other. Generally the variance decreased. The discussion as to an improvement of the AMP-manual lead to the following suggestions: In defining a psychopathological item it is important to differentiate more precisely between the patient's subjective experience and his behaviour, to find better criteria concerning the rating of different intensities and to improve discrimination between singular items. Furthermore the logic of documentation procedure should be unequivocal and more practical. Following the seminar there was a symposion discussion more general and some specific methodological problems of the further development of the AMP-system. Finally study groups were formed to elaborate on the topics "AMP-manual", "rater-training", "social data" and "electronic data processing".

Berlin↗
PubMed1975

Early and late changes in nonhistone chromatin proteins accompanying rat liver regeneration.

Chromatin was isolated from 0.025 M citric acid nuclei of regenerating rat liver at 1,5,18,24, and 48 hr posthepatectomy. The total protein to DNA ratios did not change significantly during this time period. However, 2-dimensional polyacrylamide gel electrophoresis of nonhistone proteins of "Chromatin Fraction II" revealed changes in the amounts of some protein spots. As early as 1 hr after hepatectomy, decreases in size and intensity were detected for protein spots Bp, B24, C18, and CQ, and increases were detected for protein spots CBL and C13. Late changes in size and intensity were found for protein spots BA and CN, which decreased in size and intensity 5 hr after hepatectomy. The spot densities and sizes for most of the nonhistone proteins underwent no significant changes in the course of liver regeneration. The increases and decreases observed in specific protein spots represent an ordered series of changes in a limited number of nonhistone proteins.

Animals↗