PubMed Health⌕ Search

Biomedical subjects

H Zahn

Publications and source records attributed to H Zahn.

At least 37 records · Page 2Linked to original sources
PubMed1982

[B22-D-arginine]insulin: synthesis and biological properties.

An analogue of porcine insulin which differs from the native molecule in that the amino-acid residue B22-L-arginine is replaced by its D-enantiomer has been synthesized. The [D ArgB22]B-chain was synthesized by the segment condensation method and purified as the di-S-sulfonate by ion exchange chromatoggraphy on SP-Sephadex at pH 3.5. Combination with native porcine sulfhydryl A-chain gave [DArgB22]insulin which was purified by ion exchange chromatography on SP-Sephadex at pH 4.5 with a linear NaCl gradient. The biological activity of this analogue as measured by glucose oxidation in rat epididymal adipocytes was 2%. Thymidine incorporation into DNA of human fibroblast was 16%. The immunoreactivity using antipork insulin antibody in a double antibody immunoassay was 4%. The receptor-binding affinity as measured by radioreceptor assays was 2% with cultured human fibroblasts and 1% with rat adipocytes. These results suggest that the L-configuration at B22-arginine is essential for retaining the biological, immunological and receptor-binding properties of the hormone.

Adipose Tissue↗
PubMed1982

Characterization of two anti-human parathyrin antisera.

Two anti-human parathyrin antisera were raised in sheep. These were characterized by radioimmunoassay using two commercially available bovine parathyrin preparations and one synthetic human parathyrin fragment (sequence 42-55 (42-Tyr)) for radioiodination. In addition, four synthetic human parathyrin fragments (sequences 1-34, 32-43, 434-68, 53-84), one bovine parathyrin peptide (sequence 28-48) and a human parathyrin standard from a tissue culture containing the intact hormone were utilized in a competitive inhibition assay against the two radiolabelled bovine parathyrin preparations. On column chromatography, both tracers revealed a difference in molecular weight, which is believed to be related to the extraction technique. The sequence fragment 44-68 of human parathyrin had the highest affinity for the two antisera when using the smaller molecular weight tracer and there were no qualitative changes observed in the presence of plasma. Using the higher molecular weight tracer, the addition of plasma to one of the antisera resulted in a higher affinity for the sequence fragment 1-34 of human parathyrin compared to that of intact parathyrin. Antiserum from a second sheep remained specific only for the mid-region (sequence 44-68) of the parathyrin molecule independent of the tracer used. Due to the antiserum's constant characteristic, it revealed a high reliability for the discrimination between plasma parathyrin levels in normal probands and in patients with hyperparathyroidism. Our data demonstrate that the specificity of the radioimmunoassay for human parathyrin is not exclusively dependent on the antibody source, but also on the tracer preparation and the protein content of the incubation medium.

Amino Acid Sequence↗
PubMed1982

[Synthesis of two median segments of human parathyrin (author's transl)].

Human parathyrin-(32-43)-dodecapeptide (H-His-Asn-Phe-Val-Ala-Leu-Gly-Ala-Pro-Leu-Ala-Pro-OH) and tyrosyl-[human parathyrin-(43-55)-tridecapeptide] (H-Tyr-Pro-Arg-Asp-Ala-Gly-Ser-Gln-Arg-Pro-Arg-Lys-Lys-Glu-OH) were prepared, using the strategy of segment condensation in combination with acid-labile protecting groups. The peptides will be utilized for radioimmunoassay and receptor-binding studies.

Amino Acid Sequence↗
PubMed1981

[Protein semisynthesis with the help of mixed anhydrides and enzymes: chemistry and synthesis of insulins].

Proteins play a prominent role in nature and their biosynthesis occurs via stepwise combination of amino acids. One can imitate this method in laboratory or synthesize the polypeptide chain by combining smaller preformed fragments (fragment condensation). Reversible protection of reactive groups and solubility problems arising are the most important features in this regard. Semisynthesis, i.e., coupling of amino acids or peptides to natural material may help to overcome these difficulties. The preparation of hybrid preproinsulin by mixed anhydride synthesis and the conversion of pork insulin to human insulin by enzyme-catalyzed peptide synthesis are two examples of the semisynthesis of proteins. In both cases optimal reaction conditions are essential for maximal yield of the product desired. In spite of the rapid improvement of gene technology, chemical peptide synthesis will retain its value for the preparation of biologically and pharmacologically interesting substances.

Anhydrides↗
PubMed1981

[An improved method for the combination of modified insulin chains (author's transl)].

A procedure is described in which the yields of the combination of insulin A and B chains, especially of modified chains, are increased. The chains are reduced simultaneously in 8M urea buffer at pH 8.6 with 2-mercaptoethanol. The thiol chains are not isolated but transferred to the oxidation buffer at pH 10.6 by means of gel chromatography on Sephadex G-25. During the chromatography about 50% of the tetrathiol A chain is oxidized specifically to a dithiol A chain disulphide, in which presumably the 6,11-disulphide ring is closen. Generally we observed that the combination yields of modified A or B chains are lower than the yields of combinations of unmodified A or B chains. It seems that there is a correlation between the biological activity of the analogue and the yield obtained by the recombination of the chains.

Chemical Phenomena↗
PubMed1980

Human proinsulin, VII: synthesis of two protected peptides corresponding to the sequences 1--45 and 46--96 of the prohormone.

The synthesis of two protected large peptides 1--45, and 46--86 covering the entire amino acid sequence of human proinsulin is described. Peptide 1--45, was synthesized from two intermediate fragments 1--23 and 24--45 and purified by countercurrent distribution in dimethylformamide system (K = 0.5). Peptide 46--86 was synthesized from two intermediate fragments 46--70 and 71--86 and purified by countercurrent distribution in two solvent systems, the dimethylformamide system (K = 0.06), and the toluene system (K = 0.16).

Amino Acid Sequence↗
PubMed1980

Human proinsulin, VIII: studies on the S-tritylation of reduced proinsulin, insulin A and B chains and their detritylation.

Reduced proinsulin and insulin A and B chains were selectively and quantitatively converted to hexa(S-trityl)proinsulin, tetra(S-trityl)A chain and di(S-trityl)B chain. These derivatives were used as models to investigate the quantitative removal of S-trityl groups. Amongst the various detritylation methods studied, the best procedure was found to be acidolytic cleavage with trifluoroacetic acid in the presence of thiophenol or benzylmercaptan as cation scavengers. The detritylated derivatives, upon oxidative sulphitolysis, yielded electrophoretically homogeneous hexa(S-sulphonate)proinsulin, tetra(S-sulphonate)A chain and di(S-sulphonate)B chain. The hexa(S-sulphonate)proinsulin was reduced and reoxidized to proinsulin.

Animals↗
PubMed1980

Preliminary results on the use of an antiserum to human parathyrin in a homologous radioimmunoassay.

A new antiserum (Ab Giselle) raised in sheep against extracted human parathyrin was evaluated and compared with an established antiserum (Ab S-478 VI) under several test conditions. The assay system contained an extracted 1--84 human parathyrin standard and a 1--84 bovine parathyrin tracer. The total assay time was 24 h and the main assay characteristics as follows: B0/T 0.28 +/- 0.02 and 50% intercept 553 +/- 47 U . 1(-1). The corresponding data for Ab S-478 VI were: B0/T 0.23 +/- 0.02 and 50% intercept 890 +/- 142 U . 1(-1). The normal range in 152 normocalcaemic volunteers (age range 16--67 years) was 10.6--423 U . 1(-1) (where 1 vial MRC reference preparation 75/549 for human parathyrin = 25 U), compared with 0--300 U . 1(-1) for Ab S-478 VI. With the new antiserum, differentiation between hypoparathyroid patients and those with normal function was often possible, and differentiation between normal and elevated levels, as in hyperparathyrinaemia, was very good. Correlation between Ab Giselle and Ab S-478 VI in 80 normal volunteers was positive (r = 0.450, p = 0.01) although the regression line showed that the antisera had different specificities (data for the regression line y = a + bx, a = 0.13, b = 0.55). Under the assay conditions, the association constant for Ab Giselle was 0.41 +/- 10(14) l . mol-1 in contrast to Ab S-478 VI which had a Ka for 0.53 x 10(10) l . mol-1 under identical conditions. Assays using Ab Giselle could be performed at room temperature, whereas those using Ab S-478 VI performed best at 0 degrees C. Preliminary results suggest that Ab Giselle is better for the routine assay of human parathyrin in serum than Ab S-478 VI, especially in the case of hypoparathyroid patients.

Adenoma↗
PubMed1979

[Synthesis of the fragments A1-,, A9-15 and A16-21 of ovine insulin A chain using the S-tert-butylmercapto residue for thiol protection (author's transl)].

The following paper describes the synthesis of the fragments A1-8, A9-15 and A16-21 of the ovine insulin A chain using the S-tert-butylmercapto residue for thiol protection. The synthesized fragments, which showed a good solubility in organic solvents, were partially deprotected with trifluoracetic acid and converted to the corresponding S-sulfonates quantitatively by oxidative sulfitolyses at pH 7.6.

Animals↗
PubMed1977

Introduction of new crosslinks into proteins.

Analysis of the crosslinks epsilon-(gamma-glutamyl) lysine and epsilon- (beta-aspartyl) lysine present in treated wool has been improved by modifying the enzymic digestion. Treatment of wool with either monocarboxylic acid chlorides in dimethylsulfoxide or with 1-fluoro-2,4-dinitrobenzene in the presence of acetate considerably decreased epsilon-amino groups and solubility. Since no information of interchain amide crosslinks was observed, the hypothesis of so-called self-crosslinking postulated by ZAHN has to be withdrawn. The effects of both treatments are explained in the light of new results. The reaction of wool with glutaraldehyde leads to a stabilization of the fiber. Experiments with glutaraldehyde and primary alkyl amines as model componds revealed that the cyclic form of the aldehyde gave the unstable N-alkyl-2,6-dihydroxypiperidine, which either looses water to give N-alkyldihydropyridine or condenses with 2,6-dihydroxytetrahydropyran to yield a copolyether which was isolated. According to recent publications, crosslinking of proteins by glutaraldehyde is due to the formation of quaternary pyridinium compounds.

Amino Acids↗
PubMed1977

Biotinylinsulins as potential tools for receptor studies.

The preparation of affinity columns that contain insulin attached to Sepharose in a targeted manner by way of biotin-avidin noncovalent bonds is described. Insulin was acylated selectively at the amino terminus of the B chain with the N-hydroxysuccinimido ester of biotin to form N(alpha,B1)-biotinylinsulin. The ability of this modified insulin to stimulate rat epididymal adipocytes was (mean +/- SD) 94 +/- 9.6% (P, 0.05) that of the control insulin. N(alpha,B1)-Biotinylinsulin displaced 4-hydroxyazobenzene-2'-carboxylic acid from avidin, demonstrating affinity for this protein. The formation of the N(alpha,B1)-biotinylinsulin-avidin complex was visualized by cellulose acetate electrophoresis at pH 4. N(alpha,B1)-Biotinylinsulin combined with avidin attached to Sepharose to form affinity columns in which the hormone was attached to the support by strong noncovalent bonds. The determination of the loading of avidin-Sepharose columns with biotinylinsulin was greatly facilitated by the attached biotin which provided a marker whose concentration could be assessed accurately by titration with avidin. Biotinylinsulin attached to avidin-Sepharose beads retained the ability to stimulate rat epididymal adipocytes. The activity of several samples of these beads was about 15% that of free biotinylinsulin, based on the amount of biotinylinsulin anchored to the support. The advantages of biotinylated hormones for the targeted attachment of hormones to solid supports are discussed.

Adipose Tissue↗
PubMed1976

[On the occurence of isopeptide bonds in heated milk protein (author's transl)].

Milk, milk products and individual milk proteins were subjected to different heat treatments either as powders or in aquous systems. After complete hydrolysis of the peptide bonds (alpha-amide bonds) by a system of four proteinases or peptidases, respectively, the samples were analysed for isopeptides. For this purpose, two chromatographic ion exchange systems were developed, each of which separates Nepsilon-(beta-aspartyl-)lysine (Asp Lys) as well as Nepsilon-(gamma-glutamyl-)lysine (Glu Lys) from the common amino acids. In samples, heated 24 h for at least 120 degrees C, 2-5% of the lysine residues are incorporated in Glu Lys-bonds. Under the heating conditions used in dairy practice, no isopeptide bonds were formed.

Animals↗
PubMed1976

Receptor-binding region of insulin.

X-ray analysis, circular dichroism, receptor binding and biological potencies of chemically modified insulins suggest that the conformation of the insulin molecule is critical to the formation of both the zinc insulin hexamer and the insulin-receptor complex. Results are consistent with an insulin receptor-binding region including many of the hydrophobic residues important to dimerisation in addition to more polar surface residues. There is a further possibility of formation of an antiparallel sheet structure between the insulin and receptor molecules in the complex similar to that between monomers in the insulin dimer.

Adipose Tissue↗
PubMed1976

The conformational protential of porcine proinsulin C-peptide.

Statistical analysis of protein sequences lends itself to the identification of regions with a definite inclination to adopt specific main-chain conformations. Application of the model of Chou and Fasman[1,2] to porcine proinsulin C-peptide localizes the tendency to form a helix in the segments (38 to 44) and (51 to 58). A tendency to beta turn formation is predicted for the segment (45 to 50). The realization of this conformational potential under native and various other conditions was examined by CD spectroscopy. Synthetic C-peptide as well as the synthetic fragments (33 - 40), (41 - 52), (41 - 61), (46 - 52), (46 - 61), and (53 - 61) were included in the study. These fragments provide breaks in the amino acid sequence in each of the potentially ordered regions. The strong helical tendency in the (51 - 58) segment can be activated in the fragments (41 - 61) and (46 - 61) by 1 per cent sodium dodecylsulfate, although the spectrum is not indicative of a classical alpha-helix. However, the conformation in the (51 to 58) segment should also be non-random in native C-peptide, since cleavage of the (46 - 61) fragment into the subfragments (46 - 52) and (53 - 61) causes considerable spectral effects. Cleavage of the other potentially helical region (38 to 44) between residues 40 and 41, on the other hand, is without spectral consequences. Therefore, this segment is unlikely to be helical in native C-peptide. In the coherent C-peptide, the helix formation which can be induced by sodium dodecylsulfate in the C-terminal part is apparently inhibited by interaction with the N-terminal half of the molecule. This interaction implies that the chain is folded back on itself, which is consistent with a high probability of bets turn formation in the segment (45 to 50). The CD spectra of the fragments (41 - 52) and (46 - 52), in which the beta turn could occur, are characterized by positive ellipticity about 213 nm. The correlation of the beta turn with this type of spectrum as well as its definite location are discussed, but cannot be proved solely on CD spectroscopic grounds.

Amino Acid Sequence↗