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R Deutzmann

Publications and source records attributed to R Deutzmann.

81 records · Page 5Linked to original sources
PubMed1985

Evidence for coiled-coil alpha-helical regions in the long arm of laminin.

Three new laminin fragments, E8, E9 and 25K with mol. wt. 50 000-280 000, were prepared from a limited elastase digest of laminin and from tissue extracts. They were similar with respect to their rod-like structure, a high alpha-helix content, the assembly from two chain segments and immunological cross-reactivity. Two of the fragments (E8 and E9) possess in addition globular domains which lack alpha-helices. Chemical, immunological and physical data together with sequence analysis strongly indicate that the alpha-helical segments are assembled in coiled-coil structures which are located in the rod of the long arm of laminin. These data give new insights into the overall structure of the protein.

Animals↗
PubMed1983

Cell-free translation of human liver apolipoprotein AI and AII mRNA. Processing of primary translation products.

Human liver apolipoprotein AI and A II poly(A+) mRNA has been translated in the cell-free rabbit reticulocyte lysate system. The structures of the two primary translation products of these two main protein components of human serum high-density lipoprotein (HDL) have been characterized. The products of the synthesis in vitro are preproapolipoproteins. The signal sequence (pre-sequence) of the primary translation product of human apo AI mRNA consists of 18 amino acids, that of apo AII of 17 amino acids. The cotranslational translocation into dog microsomal vesicles is associated with the cleavage of these sequences by the signal peptidase releasing the proapolipoproteins AI and AII, both extended by an N-terminal hexapeptide. Preproapolipoprotein AII is synthesized in its monomeric form consisting of 100 amino acids. Pro-apo AII is present in the vesicles of the endoplasmic reticulum also as monomer. Sequencing of the radiolabelled signal sequences of both pre-forms revealed their strongly hydrophobic nature. Despite the high affinity of HDL-apolipoproteins for complex lipids their secretion requires these hydrophobic signal sequences for translocation. Internal recognition sequences in the native apoproteins are not responsible for the transmembrane transport.

Amino Acid Sequence↗
PubMed1983

The primary structure of bovine brain myelin lipophilin (proteolipid apoprotein).

The amino-acid sequence of bovine myelin lipophilin (proteolipid apoprotein, Folch-protein) has been completed. Lipophilin is a 276 amino acid residues containing, extremely hydrophobic membrane protein with molecular mass 30,000 Da. The sequence determination was based on automated Edman degradation of four tryptophan and four cyanogen bromide fragments and of proteolytic peptides of complete lipophilin as well as the fragments obtained by chemical cleavage. Four additional sequences were determined which led to the completion of the primary structure. Lipophilin is esterified at threonine-198 by long chain fatty acids (palmitic, stearic and oleic acid). The attachment site has been established at the same threonine residue in three different peptides isolated from thermolysinolytic, papainolytic and chymotrypsinolytic hydrolysates. This threonine residue is part of a hydrophilic segment of lipophilin. The covalent fatty acyl bond is being discussed together with important structural and functional properties of this membrane protein which can be derived from sequence information. New separation and purification methods of hydrophobic and hydrophilic polypeptides for this sequence determination (fractional solubilization, silica gel exclusion, high-performance liquid chromatography) had to be elaborated as indispensable tools. They are generally applicable to the structural analysis of hydrophobic membrane proteins. Four long (26, 29, 40 and 36 residues) and one medium long (12 residues) hydrophobic segments are separated by four predominantly positively and one negatively charged hydrophilic segments. On the basis of structural data a model for the membrane integration of lipophilin is proposed.

Amino Acid Sequence↗
PubMed1982

Analysis of the primary structure of the strongly hydrophobic brain myelin proteolipid apoprotein (lipophilin). Isolation and amino acid sequence determination of proteolytic fragments.

Proteolipid aproprotein (lipophilin) and DM-20 protein from bovine brain white matter proved to be identical in polyacrylamide gel electrophoresis and automated Edman degradation of the N-terminal end over 20 cycles. Lipophilin can be hydrolysed by trypsin, thermolysis, chymotrypsin and subtilisin. We describe here a new, effective and rapid high-performance liquid chromatographic separation method for hydrophilic polypeptides according to molecular mass on an analytical and preparative scale. Three large and several small peptides have been isolated from the tryptic and thermolysinolytic hydrolysate and purified by combined molecular sieve and high-performance chromatographic separation and purification for automated Edman degradation. 40 amino acid residues of the large tryptic fragment and sequences of 43 and 22 amino acids of two thermolysinolytic fragments have been determined. These three polypeptides are partial structures of the l4 kDa large tryptophan fragment 1 or the cyanogen bromide fragment I (18-19 kDa). Thermolysin also releases a polypeptide from incompletely reductively carboxymethylated lipophilin which is cleaved into the large thermolysin fragment mentioned, 22 residues of which were analysed, and a 14 amino acids long sequence of tryptophan fragment IV, described in the previous paper. Reductively carboxymethylated liprophilin, the lysine side chains of which were blocked with maleic anhydride, can be cleaved at arginine specific sites. Bio-Gel P-150 and high-performance chromatographic purification yielded a polypeptide, which upon performic acid oxidation was split into a 15 kDa and a 7.8 kDa polypeptide. The 15 kDa polypeptide resembles the N-terminal end as proven by 31 cycles in Edman degradation. The 7.8 kDa polypeptide corresponds to the 72 amino acid C-terminal sequence, which equals cyanogen bromide fragments II, III and IV and embraces tryptophan fragment IV.

Amino Acid Sequence↗
PubMed1982

Lipophilin (proteolipid apoprotein) of brain white matter. Purification and amino acid sequence studies of the four tryptophan fragments.

The chemical cleavage of lipophilin (proteolipid apoprotein) from bovine brain white matter with HBr/dimethyl sulfoxide at the tryptophan residues, under conditions adapted to this hydrophobic protein, releases four fragments with approximate molecular masses 14 kDa (Trp I), 6.8 kDa (Trp IV), 5.2 kDa (Trp III) and 2.1 kDa (Trp II). These fragments have been separated and purified by a combination of solvent distribution, molecular sieve chromatography (Bio-Gel P-150) and high-performance liquid chromatography for automated Edman degradation and combined gas-liquid chromatography/mass spectroscopy. The complete amino acid sequences of Trp II and III and large sequences of Trp I are reported in this communication. The amino acid sequence of Trp IV and the sequences of peptides releasable from lipophilin by proteolytic enzymes (trypsin, thermolysin, subtilisin, chymotrypsin) have been described in previous reports from this laboratory. Despite two small gaps in the complete primary structure of lipophilin from myelin of central nervous system, our sequence data suggest the arrangement of four long hydrophobic sequences (30-40 apolar amino acid residues) within the hydrophobic core of the myelin lipid bilayer, linked by three hydrophilic regions at the aqueous membrane interphase. These features lend lipophilin the properties of a polytopic membrane protein.

Amino Acid Sequence↗
PubMed1982

Primary structure of the C-terminal cyanogen bromide fragments II, III and IV from bovine brain proteolipid-apoprotein.

Purified lipophilin from bovine brain white matter was reductively carboxymethylated and then cleaved by cyanogen bromide into four fragments: CNBr I 18-19 kDa, CNBr II 5 kDa, CNBr III 2.1 kDa and CNBr IV 0.7 kDa. Hydrogenbromide/dimethylsulfoxide and 3-bromo-2-(2-nitrophenylsulfenyl)skatol (BNPS-skatol) cleaved lipophilin into four fragments of molecular masses of approximately 14000 (Trp I), 2100 (Trp II), 5000 (Trp III) and 7000 Da (Trp IV). Separation and purification of the peptides for liquid phase sequenator degradation was achieved by high performance liquid chromatography. In addition proteolytic cleavage of the Trp IV fragment with trypsin facilitated the alignment of the peptides. An effective control of the sequenator data came from the partial acid hydrolysis of the Trp IV fragment, which yielded di-, tri- and tetrapeptides. The mixture was N-trifluoroacetylated, the amide (peptide) bonds and carboxyl groups were reduced with B2D6 (hexadeuterodiborane) and the polyaminoalcohols derivatized with chlorotrimethylsilane. These derivatives were separated and identified by capillary gas-liquid chromatography/mass spectrometry. Extensively overlapping sequences support the data obtained by Edman degradation in a liquid phase sequenator of the CNBr peptides II, III and IV, the 72 amino acid residues containing C-terminal sequence of lipophilin of molecular mass (7520 + x) Da, which includes the Trp IV fragment.

Amino Acid Sequence↗
PubMed1978

Quantitative ESR-studies of decay-produced radicals in 5-iododeoxyuridine labeled with 125I, 131I or tritium: role of the Auger effect.

In view of the enhanced biological damage caused by the "Auger nuclide" iodine-125, we have carried out quantitative ESR-studies of the radical formation in polycrystalline 5-iododoeoxyuridine (IUdR) resulting from the following internal or external radiation sources: (1) Decay of 3H, 125I or 131I in labeled IUdR; (2) Lanthanum K-photons corresponding to the K-edge of iodine; (3) 60Co gamma-rays. The results clearly indicate that inner shell ionization with its accompanying Auger effect as caused by the lanthanum K X-ray produces about 30% more free radicals per unit dose absorbed than 60Co gamma-rays, when considering the long-lived secondary radicals. Similarly, the concentration of free radicals is by about 30% higher in 125I- than in 131I-labeled IUdR at comparable doses. In the case of 3H-labeled IUdR the dose curve is almost identical with that observed for 125I-labeled IUdR. The results are discussed in terms of a localized radiation damage from low energy electrons.

Cobalt Radioisotopes↗
PubMedDate not supplied

Purification and properties of acridone synthase from cell suspension cultures of Ruta graveolens L.

Acridone synthase has been purified from cell suspension cultures of Ruta graveolens using a combination of gel filtration and ion exchange chromatography. The purified enzyme has an apparent molecular weight of 69 kDa on gel filtration and a subunit structure on SDS-PAGE of 40 kDa. The apparent Km-values are 10.64 microM and 32.8 microM for N-methylanthraniloyl-CoA and malonyl-CoA, respectively. Tryptic digestion of the homogeneous acridone synthase was performed. Seven of the peptides were chosen for microsequencing. The homology of the amino acid sequences from this particular polypeptide and corresponding peptides from chalcone synthase 3 from garden pea amounted to 76%.

Acyltransferases↗