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Biomedical subjects

C Nowicki

Publications and source records attributed to C Nowicki.

13 recordsLinked to original sources
PubMed1992

Presence and subcellular localization of tyrosine aminotransferase and p-hydroxyphenyllactate dehydrogenase in epimastigotes of Trypanosoma cruzi.

Cell-free extracts of epimastigotes of Trypanosoma cruzi contain tyrosine aminotransferase (TAT) and p-hydroxyphenyllactate dehydrogenase (pHPLDH). The TAT activity could be separated from aspartate aminotransferase (ASAT) by polyacrylamide gel electrophoresis or DEAE-cellulose chromatography; the latter procedure also allowed complete separation of pHPLDH. The subcellular localization of both T. cruzi enzymes, as determined by digitonin extraction, subcellular fractionation by differential centrifugation, and isopycnic ultracentrifugation in sucrose gradients, was mainly cytosolic, with low mitochondrial activities.

Animals↗
PubMed1991

Chemical modification of human growth hormone with N-acetylimidazole. Effect on binding capacity to lactogenic and somatogenic receptors.

The effect of acetylation of tyrosine residues on the binding capacity of human growth hormone (hGH) to rat liver lactogenic and somatogenic receptors was studied. When 3.7 tyrosine and 4.8 lysine residues were acetylated with N-acetylimidazole, both the in vivo and the in vitro capacities of hGH to compete with 125I-labeled bovine growth hormone for somatogenic binding sites greatly decreased. Acetylation also affected the in vitro binding capacity to lactogenic sites. Most of the somatogenic binding activity was recovered by hydroxylamine treatment, which removes O-acetyl groups from tyrosine residues but not N-acetyl groups from lysine residues. The same treatment partially restored lactogenic binding capacity. The reactivity of hGH tyrosine residues to N-acetylimidazole, together with previous evidence, suggests that: (a) Tyrosine residues 160 and 164, when acetylated, are likely to be responsible for the low binding activity of acetylated hGH. (b) Tyrosine 160 may play a significant role in hGH interaction with lactogenic receptors.

Acetylation↗
PubMed1990

Modification of arginine residues in human growth hormone by 1,2-cyclohexanedione: effects on the binding capacity to lactogenic and somatogenic receptors.

Reactivity of arginine residues in human growth hormone was studied by reaction with 1,2-cyclohexanedione. Kinetic analysis of the data showed a good fit to a pseudo first order curve, with an apparent velocity constant k = 1.26 x 10(-2) min-1 and a maximum modification of 9.6 out of the 11 arginines of the molecule. Modification led to a decrease in binding capacity to both lactogenic and somatogenic rat liver receptors. In either case Tsou plots suggest that the modification of two arginine residues is responsible for this behavior, although it cannot be ascertained whether the two relevant residues are the same for both receptor types. Circular dichroism studies indicated no apparent changes in protein conformation in the modified hormone. Binding capacity was restored upon regeneration of arginines by incubation with Tris-HCl buffer. Only the carboxy-terminal peptide was isolated by HPLC from a tryptic digest of succinylated Arg-modified hGH, indicating that 183 is the nonreacting arginine residue.

Animals↗
PubMed1988

An approach to the three-dimensional structure of bovine growth hormone based on chemical modification and secondary structure prediction.

Three different methods have been applied to the prediction of secondary structure. The prediction that better fitted the chemical data was chosen. Two regions of the bovine growth hormone molecule (111-117 and 166-174) appear to be exposed to the solvent, according to hydropathic analysis but have several charged residues not reactive towards their specific reagents. Two molecular domains are postulated, each one bearing a region with charged residues on its surface and interacting with the other in the molecule by means of saline bridges. The hydrophobic core of the molecule is formed by the ensemble of the hydrophobic region predicted between residues 81 and 108, and the hydrophobic faces of the amphiphilic helices 109-127 and 9-33.

Amino Acid Sequence↗
PubMed1988

Ovine prolactin and human growth hormone derivatives. Specific modification of their alpha-amino groups.

The alpha-amino group of ovine prolactin (oPRL) and human growth hormone (hGH) was selectively modified by transamination with glyoxylic acid. No difference was found in the binding capacity of transaminated oPRL to rat liver lactogenic receptors with respect to its control, although both samples showed a decrease in its binding capacity with reference to the native hormone. This decrease was due to conformational changes caused by the reaction conditions and not by the transamination itself, as shown by the circular dichroism spectra. Transaminated hGH retained the full binding capacity of the hormone. These results suggest that the alpha-amino group is not relevant for the binding to lactogenic liver receptors in both lactogenic hormones.

Animals↗
PubMed1987

Identification of intramolecular crosslinks in bovine growth hormone after two-step modification with 1,5-difluoro-2,4-dinitrobenzene.

Derivatives of bovine growth hormone, containing monoaminotyrosyl residues in positions 35, 42 and 174, were treated at pH 3.6 with a bifunctional reagent, 1,5-difluoro-2,4-dinitrobenzene. Under these conditions aminotyrosyl groups reacted. On changing the pH to 9.3, the second fluorine atom of the reagent was substituted with the sterically adjacent side groups of lysine, since the excess of reagent had been previously removed. The modified protein underwent cyanogen bromide treatment. Peptides containing the crosslinks were purified from tryptic digests of the cyanogen bromide fragments by HPLC. Results show that aminoTyr 174 was able to form dinitrophenylene bridges with Lys 111, Lys 29 and Lys 170. AminoTry 35 was found crosslinked to Lys 29. Taking into account the size of the reagent, it may be inferred that Lys 29, 111 and 170 are located at approximately 5 A from Tyr 174 in the bovine growth hormone molecule.

Amino Acid Sequence↗
PubMed1985

Evidence for the steric proximity of Tyr 174 and Lys 111 in bovine growth hormone.

Bovine growth hormone was modified by reaction with 1,5-difluoro-2,4-dinitrobenzene under conditions favouring production of intramolecularly crosslinked derivatives from monomeric molecules. The monomeric fraction, isolated by chromatography on Sephadex G-100, was oxidized or reduced and carbamidomethylated and trypsin digested. The resulting peptides were fractionated on SP-Sephadex and further purified by peptide mapping or HPLC. Two modified peptides containing sequences 108-112 or 108-113 and 171-176 of bGH were obtained, including a dinitrophenylene bridge between lysine 111 and tyrosine 174, thus suggesting the stereochemical proximity of these residues.

Amino Acid Sequence↗
PubMed1984

Hydrophobic residues involved in the interaction between protomers of the bovine growth hormone dimer. Methionine and tyrosine residues.

The bovine growth hormone dimeric form covalently stabilized by cross-linking with dimethyl suberimidate (DMS) and the hormone modified by DMS without forming covalent links with other hormone molecules (DMS-bGH) were oxidized with chloramine-T at molar-ratios of 2 and 50 with respect to methionine content. The extent of oxidation undergone by each methionine residue, estimated on the purified tryptic peptides, closely resembled that obtained for the native hormone, thus suggesting that methionine residues are not involved in the protomers interaction area. Evaluation of the reactivity of tyrosine residues toward tetranitromethane indicated that, in both the covalent dimer and DMS-bGH, tyrosine residues 35, 174 and 142 are the more susceptible to undergo reaction. Net charges can be induced in the iodotyrosine residues in the iodinated hormone, by setting the pH at 10.5. At this pH, dissociation of a fraction of uniformly iodinated hormone was observed in the derivatives containing 2 or more iodine atoms, indicating that tyrosine residues might integrate the contact area between protomers.

Chloramines↗
PubMed1981

Modification of lysine 69 reactivity in bovine growth hormone by carbamylation of its N-terminal group.

Bovine growth hormone was carbamylated under conditions that assure full reaction of the N-terminal residue. Approximately 28% of lysine 179 and 7% of lysine 143 were also carbamylated. The modified hormone retained an important growth promoting activity and was as effective as the native hormone in competition assays in vivo for the receptors in rat liver. However, a change in its conformation must occur since lysine 69, which is resistant to trinitrophenylation in the native hormone, reacted easily and under mild conditions, in the carbamylated protein, The growth promoting activity and binding properties of the carbamylated and trinitrophenylated hormone were practically nil.

Animals↗
PubMed1979

Antibodies against haptens in the study of structure and biological activity of growth hormone.

The alpha-amino group of bovine Growth Hormone was selectively modified with TNBS with no detectable changes in growth promoting activity. TNP was used as hapten for the production of antibodies against the end terminal region of bGH. The region near the alpha-amino group is not involved in the binding of bGH to rat liver cells, and it seems to be away from the part of the molecule that interacts with the cell binding sites.

Amino Acids↗