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Biomedical subjects

K D Gibson

Publications and source records attributed to K D Gibson.

At least 55 records · Page 3Linked to original sources
PubMed1977

The effect of beta-D-xylosides on chondroitin sulphate biosynthesis in embryonic chicken cartilage in the absence of protein synthesis inhibitors.

Incorporation of [35S]]sulphate, [3H]glucose and [3H]serine into glycosaminoglycans and proteoglycans of embryonic-chicken sternum was measured in vitro in incubation medium containing 4-methylumbelliferyl beta-D-xyloside or p-nitrophenyl beta-D-xyloside at low concentrations, and in the absence of inhibitors of protein synthesis. Incorporation of sulphate was decreased by 80% in incubations in which 1mM-4-methylumbelliferyl beta-xyloside or 2.5 mM-p-nitrophenyl beta-xyloside was present; under these conditions, serum factors stimulated incorporation to only a small extent. When the concentration of the xyloside was decreased tenfold, incorporation of sulphate was inhibited by 60-70%, but when normal human serum or L-3,3',5-tri-iodothyronine or both were also added to the incubation medium, incorporation was markedly stimulated. Experiments in which [35S]sulphate and [3H]glucose were incorporated simultaneously, and enzymic analysis of glycosaminoglycans formed in such experiments, indicated that chondroitin sulphate formed in the presence of 0.1 mM-4-methylumbelliferyl beta-xyloside contained 30-40% less sulphate than did chondrotin sulphate synthesized in the absence of xylosides. Similar experiments, with [3H]serine instead of [3H]glucose, suggested also a 20-30% decrease in chain length of the chondroitin sulphate; this was confirmed by direct gel filtration of labelled glycosaminoglycans on a calibrated column. Incorporation of [3H]glucose or [3H]serine was stimulated by serum and tri-iodothyronine in parallel with incorporation of sulphate. The changes seen in the total chondroitin sulphate were mirrored in the major proteoglycan fraction, purified by isopycnic centrifugation of salt-extracted proteoglycans. The labelling pattern of chondroitin sulphate from this proteoglycan indicated that decreased sulphation of chondroitin sulphate was largely due to the inferior ability of short polysaccharide chains to accept sulphate, with some direct interference with transfer of sulphate to all chains. The results also suggested that the action of serum factors on synthesis of proteochondroitin sulphate is exercised at the level of either protein synthesis or transport to the sites of initiation of polysaccharide synthesis.

Animals↗
PubMed1976

Effects of medium composition and metabolic inhibitors on glycosaminoglycan synthesis in chick embryo cartilage and its stimulation by serum and triiodothyronine.

Incorporation of inorganic sulfate into glycosaminoglycans of chick embryo sternum is stimulated by serum and triiodothyronine. Variations in the amino acid content of the medium, and in particular in the concentration of glutamine, changed the incorportion in control and stimulated sterna to the same degree. Omission of Na+ from the medium greatly reduced incorporation in both control and stimulated sterna; incorporation, and its stimulation by triiodothyronine, was restored by raising the concentration of Na+. Ouabain and valinomycin inhibited incorporation more than 90%, and triiodothyronine did not stimulate under these conditions. Puromycin and cycloheximide also inhibited incorporation almost completely, and abolished the stimulation by triiodothyronine and serum. Addition of p-nitrophenyl-beta-xyloside, in the presence of of puromycin ir cycloheximide, restored sulfation to a level of 5-10% of the control value; however, this level of incorporation was not increased by addition of serum or triiodothyronine. Actinomycin D, colchicine and vinblastine inhibited incorporation by 40% or less at the highest concentrations tested; however, these three agents completely abolished the ability of triiodothyronine to stimulate incorporation. Lumicolchicine and cytochalasin B decreased incorporation in controls slightly but did not affect the stimulation by serum or triiodothyronine. The results indicate that thyroid hormones stimulate glycosaminoglycan synthesis only under conditions which support efficient synthesis in control incubations, and suggest that microtubule formation may be essential to the mode of action of thyroid hormones in this system.

Amino Acids↗
PubMed1976

Stimulation of proteoglycan synthesis in chick embryo sternum by serum and L-3,5,3'-triiodothyronine.

Incorporation of sulfate into alcian blue-precipitable glycosaminoglycan of 12-day-old chick embryo sterna is stimulated by addition, separately or together, of normal human serum and physiological concentrations of thyroid hormones (Audhya, T.K., and Gibson, K.D. (1975) Proc. Natl. Acad, Sci. U. S. A. 72, 604--608). We present evidence that this stimulation is due to increased synthesis of at least one proteoglycan, with minor alterations in the size and chemical composition of the glycosaminoglycans. Pulse-chase experiments showed no detectable loss of label during the chase, in control sterna or sterna incubated with serum and L-3,5,3'-triiodothyronine; thus, all incorporation was the result of synthesis of glycosaminoglycans. In double-label experiments, with 35SO4(2-) and [3H]acetate, the molar ratio of 3H and 35S incorporated into glycosaminoglycans was changed little, if at all, by addition of serum or triiodothyronine or both, at concentrations which increased incorporation up to 2-fold. Glycosaminoglycans isolated from these and other incubations gave similar elution patterns from agarose columns, and identical electrophoretic patterns on cellulose acetate. Digestion with chondroitinase ABC (chondroitin ABC lyase; EC 4.2.2.4.) showed that incorporation was into chondroitin sulfate and possibly hyaluronic acid, and that the proportions of non-sulfated, 4-sulfated, and 6-sulfated disaccharide units differed little between stimulated and unstimulated sterna. Incorporation of [3H]serine into glycosaminoglycans from papain digest of sterna paralleled incorporation of 35SO4(2-), and indicated a number average molecular weight between 21,000 and 25,000 for the newly synthesized chondroitin sulfate. This value was confirmed by gel filtration chromatography, which also showed that the average molecular weight of the newly synthesized chondroitin sulfate decreased up to 15% under conditions of 2-fold stimulation. Proteoglycans were extracted from sterna incubated with [3H]serine and 35SO4(2-) and analyzed by isopycinic centrifugation in CsCl and by zone sedimentation in a sucrose gradient. A major proteoglycan fraction could be separated by either method. Incorporation of both isotopes into this proteoglycan fraction, and into glycosaminoglycans isolated after papain digestion, was stimulated in a coordinate manner. Almost identical results were obtained with both separation techniques. The results indicate that the synthesis of the major proteoglycan, and probably also of a minor one, is stimulated by serum and triiodothyronine.

Acetates↗
PubMed1976

Nonsuppressible insulin-like activity and thyroid hormones: major pituitary-dependent sulfation factors for chick embryo cartilage.

Serum from hypothyroid hypophysectomized rats did not stimulate sulfation or incorporation of amino acids into chick embryo sterna. When such rats were treated for a short time with growth hormone (somatotropin), their serum stimulated incorporation both of sulfate and of amino acids. The different actions of the two types of sera were not due to changes in thyroid state. The results support the existence in serum of a sulfation factor for chick embryo cartilage that is dependent upon growth hormone. Highly purified preparations of nonsuppressible insulin-like activity from human serum stimulated incorporation of amino acids, and of uridine into RNA, in chick embryo sterna in vitro; chondrocytes prepared from this tissue had specific high-affinity binding sites for this insulin-like activity. However, sulfate incorporation was stimulated very little, unless serum from hypothyroid hypophysectomized rats was also present. When L-3,5,3'-triiodothyronine was added as well, the stimulation was enhanced further. From these and other experiments, we conclude that (i) nonsuppressible insulin-like activity or a closely related peptide is the growth-hormone-dependent growth and sulfation factor for chick embryo cartilage: (ii) a second, unidentified factor must be present for the insulin-like activity to stimulate sulfation; and (iii) stimulation of sulfation by thyroid hormones in vitro is additive to that of nonsuppressible insulin-like activity.

Animals↗
PubMed1975

Enhancement of somatomedin titers of normal and hypopituitary sera by addition of L-triiodothyronone in vitro at physiological concentrations.

Somatomedin potencies of sera were assayed by following sulfation of mucopolysaccharides in chick embryo sterna in vitro. Apparent potencies of sera from hypophysectomized rats, maintained on a low-iodine diet, were restored to levels above normal by addition to the incubation medium of L-triiodothyronine at 10-7 mol/liter of serum. The potencies of normal rat, human, and fetal calf sera were raised 1.3- to 3-fold by addition of triiodothyronine at 10-9-10-7 mol/liter of serum. L-Thyroxine was about 10 times less effective than triiodothyronine. Triiodothyronine alone did not stimulate sulfation to nearly the same extent as triiodothyronine plus serum, even at higher concentrations. Serum could not be substituted in this system by any of six purified hormones, nor by trace metals or amino acids not included in the incubation mixture. It is concluded that triiodothyronine, in combination with a factor in serum, causes a rapid stimulation of sulfation in chick embryo cartilage, and that thyroid hormones may be involved in the action of normal serum on this tissue.

Animals↗
PubMed1972

Changes in ribonucleic acid turnover during aerobic and anaerobic growth in Rhodopseudomonas spheroides.

Differences in the rate and extent of degradation of ribonucleic acid (RNA) labeled by a 30-sec pulse in aerobically or anaerobically grown Rhodopseudomonas spheroides have been studied by using rifampin to block RNA synthesis. In anaerobic cultures, unstable RNA is degraded with a half-life of 1.25 to 2.0 min, and about 40% of the pulse-labeled RNA is stable. In aerobic cultures, the half-life of unstable RNA is increased to 2.5 to 4.0 min, and 50% of the RNA is stable. When aerobic cultures are transferred to anaerobic conditions, there is a rapid drop in half-life and in the proportion of stable RNA. When anaerobic cultures are made aerobic, the reverse changes occur after a lag of about 30 min. Addition of puromycin to either aerobic or anaerobic cultures caused the pulse-labeled RNA to be degraded at the same rate and to the same extent as the RNA in an anaerobic control culture. In contrast, addition of chloramphenicol enhanced the difference in RNA half-life and increased the proportion of stable RNA by about 10% in each case. It is concluded that there is a difference in the stability of an RNA component under aerobic and anaerobic conditions.

Aerobiosis↗
PubMed1972

Ribonucleic acid from aerobically and anaerobically grown Rhodopseudomonas spheroides: comparison by hybridization to chromosomal and satellite deoxyribonucleic acid.

Ribonucleic acid (RNA) species from aerobically and anaerobically grown Rhodopseudomonas spheroides were compared via hybridization to deoxyribonucleic acid (DNA). Both long-labeled and stable RNA bound to chromosomal DNA to the same extent, regardless of derivation. About 4% of the chromosomal DNA hybridized with total cell RNA and about 0.08% with stable RNA. About 4% of the mixed satellite DNA could be hybridized to total cell RNA from aerobic or anaerobic cultures, whereas essentially no stable RNA formed a hybrid with this DNA. Hybridization competition experiments with aerobic and anaerobic pulse-labeled RNA and chromosomal or satellite DNA demonstrated that no qualitative differences existed between the RNA species. It is concluded that identical species of RNA in the same relative amounts are synthesized by R. spheroides during aerobic or anaerobic growth on the same medium.

Aerobiosis↗
PubMed1971

Deficiencies of chromatophore proteins in some mutants of Rhodopseudomonas spheroides with altered carotenoids.

Chromatophore proteins of a wild type and three mutant strains of Rhodopseudomonas spheroides were examined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The mutants consisted of a green and a blue-green one, whose phenotypes were essentially the same as those of known mutants, and a brown one, which may be a double mutant and represents a new phenotype. Wild-type chromatophores contained at least six major and seven minor protein bands, with molecular weights ranging from 10,000 to 65,000. The green mutant contained the same protein bands in the same relative quantities. The brown mutant had one protein completely missing and no other alterations. The blue-green mutant was deficient in a different protein, and had reduced quantities of all proteins with molecular weights less than 25,000. Chromatophores were separated into a fraction containing the reaction centers and a fraction containing the light-harvesting bacteriochlorophyll by treatment with sodium dodecyl sulfate. Eight of the proteins were found only in the reaction center fraction, one was only in the light-harvesting fraction, and the remainder were present in both fractions. The protein missing from the brown mutant was found to be a component of the reaction center fraction, whereas the proteins which were missing from the blue-green mutant were all components of the light-harvesting fraction. Some implications for the structure and biogenesis of chromatophores are discussed.

Acrylates↗
PubMed1969

Minimization of polypeptide energy, vi. Systematic searches for low-energy conformations of deca-L-alanine and the octapeptide loop of ribonuclease.

A succession of apparent local energy minima, in which the energies decrease successively, have been located for deca-L-alanine and the octapeptide loop of ribonuclease. The lowest energy minima were approximately 40 kcal/mole lower than the highest ones. The results suggest that the computational methods used here might be of considerable use in searching for the global energy minimum of larger polypeptides.

Journal Article↗
PubMed1969

Minimization of polypeptide energy. VII. Second derivatives and statistical weights of energy minima for deca-L-alanine.

The matrix of second derivatives of 21 apparent local energy minima of deca-L-alanine has been computed. The results show that all these conformations are true local energy minima and, therefore, potentially stable conformations for this peptide. Calculations of the relative statistical weights of these conformations confirm earlier theoretical considerations on the importance of the librational free energy of stable conformations of peptides. As predicted earlier, the relative statistical weights of the local energy minima do not always fall in the same order as the relative energies.

Alanine↗