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Biomedical subjects

H Welfle

Publications and source records attributed to H Welfle.

At least 55 records · Page 3Linked to original sources
PubMed1986

Comparison of the structure of ribosomal 5S RNA from E. coli and from rat liver using X-ray scattering and dynamic light scattering.

The structure of eukaryotic ribosomal 5S RNA from rat liver and of prokaryotic 5S RNA from E. coli (A-conformer) have been investigated by scattering methods. For both molecules, a molar mass of 44,500 +/- 4,000 was determined from small angle X-ray scattering as well as from dynamic light scattering. The shape parameters of the two rRNAs, volume Vc, surface Oc, radius of gyration Rs, maximum dimension of the molecule L, thickness D, and cross section radius of gyration Rsq, agree within the experimental error limits. The mean values are Vc = 57 +/- 3 nm3, Oc = 165 +/- 10 nm2, Rs = 3.37 +/- 0.05 nm, L = 10.8 +/- 0.7 nm, D = 1.57 +/- 0.07 nm, Rsq = 0.92 +/- 0.01 nm. Identical structures for the E. coli 5S rRNA and the rat liver 5S rRNA at a resolution of 1 nm can be deduced from this agreement and from the comparison of experimental X-ray scattering curves and of experimental electron distance distribution functions. The flat shape model derived for prokaryotic and eukaryotic 5S rRNA shows a compact region and two protruding arms. Double helical stems are eleven-fold helices with a mean base pair distance of 0.28 nm. Combining the shape information obtained from X-ray scattering with the information about the frictional behaviour of the molecules, deduced from the diffusion coefficients D020, w = (5.9 +/- 0.2) X 10(-7) cm2 s-1 and (6.2 +/- 0.2) X 10(-7) cm2 s-1 for rat liver 5S rRNA and E. coli 5S rRNA, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1985

Protein--RNA interaction in the rat liver 5S rRNA-protein L5 complex studied by digestion with ribonucleases.

Protein-RNA interactions in the 5S rRNA-protein L5 complex from rat liver ribosomes were studied by limited digestion of free and protein bound 5S rRNA with ribonuclease A and T1. In the complex with protein L5 the digestion of 5S rRNA by ribonuclease T1 is decreased at G37 and G89, whereas U38 and C39, and to a lower extent also C10 and U12 become accessible for ribonuclease A.

Animals↗
PubMed1985

A-form to A'-form conformational switch of double helices in rat liver 5S and 5.8S rRNA. Solution X-ray scattering evidence and circular dichroic measurements.

The wide-angle X-ray scattering of rat liver 5S rRNA and 5.8S rRNA molecules showed significant differences in the positions of the scattering maxima when dissolved in Mg2+-containing Tris/HCl buffer or in Mg2+-depleted buffer. A comparison of the experimental curves with theoretical curves calculated from atomic coordinates of double-helical models proved a switch from A form to A' form of the double-helical regions within the molecules by changing the buffer conditions. This result was supported by circular dichroic measurements. The A to A' transition may have important consequences for RNA-protein interactions.

Animals↗
PubMed1985

Conserved unpaired adenine residues are important for ordered structures of 5S ribosomal RNA. An infrared study of the secondary and tertiary structure of Thermus thermophilus 5S rRNA.

An improved set of infrared calibration spectra for the determination of G X C and A X U base pairs leads to 32 +/- 3 G X C (+ G X U) and 4 +/- 1 A X U base pairs for Thermus thermophilus 5S RNA in the presence and absence of Mg2+. These results give further support for the consensus secondary structure of 5S RNA recently proposed by several groups. T. thermophilus 5S RNA shows, in the presence of Mg2+, a distinct two-step thermal melting of its ordered structure. Based on new data about the stacking dependence of infrared intensities of unpaired ribonucleotides the spectral changes of the low-temperature transition should be explained by melting of stacked arrangements of unpaired bases and/or non-standard base pairs. Striking is the reduction in A stacking, which is not related to the melting of A X U base pairs, indicating the importance of the mostly conserved unpaired adenines for the Mg2+ stabilized higher-order structures especially within internal loops of 5S RNA.

Adenine↗
PubMed1983

Laser Raman studies of the 5 S rRNA-protein L5 complex of rat liver ribosomes.

The effects of ribosomal protein L5 on the conformation of 5 S rRNA in the 5 S rRNA-protein L5 complex extracted from rat liver ribosomes have been studied by laser Raman spectroscopy. A comparison of the spectra shows small protein-induced conformational changes in the 5 S rRNA, but most of the base-paired regions appear to be present in the complex with protein L5 as well as in the free 5 S rRNA. Furthermore specific interactions between 5 S rRNA and protein L5 are indicated. Cytosine (and/or uracil) residues in single-stranded regions and the N(7) of guanine are engaged in interactions with the protein as suggested by the Raman data.

Animals↗
PubMed1982

Two distinct conformations of rat liver ribosomal 5S RNA.

Three different conformers of rat liver 5S ribosomal RNA were investigated by partial nuclease cleavage technique using S1 nuclease and cobra venom endoribonuclease (CVE) as conformational probes. Urea-treated and renatured 5S RNA co-migrate on non-denaturing gels, but exhibit distinct differences in their nuclease cleavage patterns. The most prominent differences in S1 nuclease and CVE accessibility of these conformers are located in region 30-50 and around nucleotides 70 and 90. The third form of 5S RNA with higher electrophoretic mobility was generated by EDTA treatment. The cleavage patterns of this 5S RNA conformer are similar to that characteristic for the renatured 5S RNA. The results demonstrate the difference in secondary structure and possibly different tertiary base-pairing interactions of 5S RNA conformers.

Animals↗
PubMed1982

Universal structural features of prokaryotic and eukaryotic ribosomal 5S RNA derived from comparative analysis of their sequences.

An extensive comparative analysis of more than fifty available sequences of ribosomal 5S RNA has been made. Both for prokaryotic and eukaryotic 5S RNA a generalized secondary structure is presented which is similar to that suggested by Nishikawa and Takemura modified in few positions only. Both generalized secondary structures contain five main helical regions and a high base-pairing content of about 65 +/- 5%. The general structural architecture of prokaryotic and eukaryotic 5S RNA molecules appears to be very similar with minor modifications within particular subgroups of organisms. Conserved and semiconserved nucleotides are accumulated in the single stranded parts of 5S RNA. Functional importance was suggested for some of these regions; other short conserved nucleotide stretches may be involved in the folding of 5S RNA molecules. In particular, we propose a tertiary base-pairing interaction between the universal invariant GUA sequence (positions 76-78 and 75-77 in prokaryotic and eukaryotic 5S RNA, respectively) and the complementary conserved CPuU sequence (positions 38-40 and 36-38) in a parallel manner. A molecular model of the 5S RNA of human KB cells was constructed, which verifies the proposed tertiary interaction, probably stabilizing the two neighboured helices E and D and a stacking arrangement of the bases in the sequence positions 67-108 (and 70-106) in eukaryotic (and prokaryotic) 5S RNAs, respectively.

Animals↗
PubMed1981

A small-angle and wide-angle x-ray scattering study of the shape and secondary structure of native 5 S RNA from rat liver ribosomes.

Native 5 S RNA from rat liver ribosomes was investigated by means of small-angle and wide-angle X-ray scattering. The radius of gyration, Rs of the molecule is 3.1 nm, the maximum dimension, L, 10.5 nm and the shape volume, Vc, about 60 nm3. The overall shape of the molecule as derived from these parameters is a flat elliptical cylinder with dimensions of 2a = 10.45 nm, 2b = 6.94 nm, H = 1.06 nm. The cross-section radius of gyration of 0.92 nm and the mass per unit length of 2610 mol X nm-1 of the 5 S RNA molecule are typical for a double helical organized RNA molecule. From wide-angle scattering data it can be concluded that the double helical regions are in the A-form characterized by an 11-fold helix with a turn angle of 32.7 degrees and a distance of 0.28 nm between adjacent base pairs. A refined electron density model of a distorted L shape is proposed for the 5 S RNA molecule.

Animals↗
PubMed1980

Physicochemical studies of the 7 S complex of rat liver ribosomes and its components.

The 7 S complex of rat liver ribosomes contains 5 S RNA and one protein (L-5). The molecular weights of these two components are 40 600 and 37 200, respectively, as estimated by analytical ultracentrifugation. Th complex stability under different solvent conditions was analyzed from the point molecular weights using a sedimentation equilibrium technique and computer stimulation. High concentrations of KCl and urea induce partial dissociation, whereas MgCl stabilizes the complex.

Animals↗
PubMed1978

Preparative two-dimensional polyacrylamide gel electrophoresis of rat liver ribosomal proteins and determination of their amino acid compositions.

1. By enlarging the dimensions of the gels used in the usual analytical two-dimensional polyacrylamide gel electrophoresis it is possible to separate much larger amounts of ribosomal protein in comparison to analytical separations. 15 mg of protein mixture of small or large subunits of rat liver ribosomes can be separated by this procedure. 2. The positions of the proteins in the two-dimensional patterns are identified with a special staining procedure. The proteins are eluted from the gels with SDS/phosphate buffers. 3. The purity of the extracted proteins was tested by one-dimensional SDS-polyacrylamide gel electrophoresis and two-dimensional electrophoresis, respectively. 24 proteins of the small and 24 proteins of the large ribosomal subunit were isolated in pure form. 4. The amino acid compositions of 24 proteins of the small and of 19 proteins of the large subunit were determined.

Amino Acids↗
PubMed1978

Number and molecular weights of the basic proteins of rat liver ribosomes.

1) Efficient separation of the proteins from rat liver ribosomes can by achieved by two-dimensional polyacrylamide gel electrophoresis. Complete separation of all components, however, is not possible with one system only. Comparison of the results obtained with different systems suggests further heterogeneity of S15, L22, L28, L33 and L35 and enables identification of S15a, S15b, L22a, L22b, L28a, L28b, L33, L33a, L35a and L36b. 2) Ribosomal proteins were substituted with iodoacetamide prior to electrophoresis or handled in all steps of the procedure in the presence of reducing agents. These procedures prevent the formation of oxidation products described erroneously as ribosomal proteins S5, S6, L15, L17 and L32 in earlier papers. 3) Estimation of the molecular weights was performed by two-dimensional separation of the small and large subunit proteins using sodium dodecyl sulphate in the second dimension. The positions of the 70 basic proteins in the 2-D patterns were identified. 4) The small and large subunit proteins have molecular weights in the range of 8000 to 35,000 and 11,000 to 55,500 Dalton, respectively. The number average molecular weights for the small and large subunit proteins are 22,500 and 26,500 Dalton, respectively. The sum of the molecular weights is 0.67 x 10(6) Dalton for the proteins of the small subunit and 1.05 x 10(6) Dalton for the proteins of the large subunit.

Acetates↗
PubMed1977

Presence of an endopeptidase activity in rat liver ribosomes.

Preparation of ribosomes using different procedures (treatment of postmitochondrial-postlysosomal supernatant or microsomes with 1% triton in 0.15 or 0.5 M KCl and subsequent sucrose gradient centrifugation; treatment of microsomes with 1.5% deoxycholate/2% triton) results in purified ribosomes which contain an endopeptidase activity detectable by breakdown of ribosomal proteins to trichloroacetic acid soluble split products. The proteolytic activity can be recovered also in the extracted proteins of whole ribosomes. With ribosomes the pH optimum of proteolytic breakdown is at about 7. The inhibition of the activity by leupeptin, DIFP and soya bean trypsin inhibitor suggests a serine type of the proteolytic activity.

Animals↗