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Biomedical subjects

J C Wallace

Publications and source records attributed to J C Wallace.

At least 145 records · Page 8Linked to original sources
PubMed1989

A key functional role for the insulin-like growth factor 1 N-terminal pentapeptide.

In order to elucidate the role of the N-terminus of insulin-like growth factor 1 (IGF-1) with respect to its biological properties, we chemically synthesized analogues of IGF-1 truncated by one to five amino acid residues from the N-terminus. In a bioassay that measured the stimulation of protein synthesis in rat L6 myoblasts, the concentrations required to produce a half-maximal response were: IGF-1, 13 ng/ml; des-(1)-IGF-1, 10 ng/ml; des-(1-2)-IGF-1, 13 ng/ml; des-(1-3)-IGF-1, 1.5 ng/ml; des-(1-4)-IGF-1, 5.1 ng/ml; des-(1-5)-IGF-1, 1200 ng/ml. When tested for their abilities to compete with 125I-IGF-1 binding to L6 myoblasts at 3 degrees C, the concentrations required for 50% competition were: IGF-1, des-(1)-IGF-1 and des-(1-2)-IGF-1, 20 ng/ml; des-(1-3)-IGF-1, 14 ng/ml; des-(1-4)-IGF-1, 40 ng/ml; des-(1-5)-IGF-1, greater than 1000 ng/ml. Receptor-binding experiments at 25 degrees C, however, gave results suggesting that the myoblasts were secreting a binding protein selective for the three longest peptides. This interpretation was confirmed by binding studies with medium conditioned by the L6 myoblasts as well as binding protein purified from MDBK-cell-conditioned medium. In both cases IGF-1, des-(1)-IGF-1 and des-(1-2)-IGF-1 competed for tracer IGF-1 binding at least 60-fold better than did the three shorter peptides. The results obtained account for the increased potency of des-(1-3)-IGF-1 and des-(1-4)-IGF-1, since their activities are not attenuated by the binding protein, and the relatively lower potency of des-(1-4)-IGF-1 is a consequence of this peptide binding less well to the L6-myoblast receptor.

Animals↗
PubMed1989

Insulin-like growth factor (IGF)-binding proteins inhibit the biological activities of IGF-1 and IGF-2 but not des-(1-3)-IGF-1.

(1) Many cell types secrete insulin-like growth factor (IGF)-binding proteins that can be expected to sequester free IGF and modify the biological activities of the growth factors. (2) A binding protein purified from bovine kidney (MDBK) cells potently inhibited the ability of IGF-2 to stimulate DNA synthesis or protein accumulation as well as to reduce rates of protein breakdown in chick embryo fibroblasts. The binding protein did not influence the biological activities of des-(1-3)-IGF-1, while effects on IGF-1 were intermediate. Since the chick embryo fibroblasts contain only the type 1 IGF receptor, the MDBK-cell binding protein must have reduced the accessibility of IGF-2 and IGF-1 to that receptor. Binding to the type 2 receptor on L6 myoblasts was also inhibited. (3) Inhibiting effects on both protein breakdown responsiveness to IGF and IGF binding to cell receptors were also observed with human amniotic fluid binding protein, although here IGF-1 and IGF-2 were equipotent. These results contrast with stimulatory responses on different IGF-1 actions of the same binding protein reported previously [Elgin, Busby & Clemmons (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 3254-3258]. (4) The biological potencies of IGF-1, IGF-2 and des-(1-3)-IGF-1 correlate inversely with their binding to proteins released into the medium by cells, so that the enhanced potency of des-(1-3)-IGF-1 is a consequence of it not binding to purified binding proteins or those released by cultured cells.

Animals↗
PubMed1989

Is dietary epidermal growth factor absorbed by premature human infants?

Urinary epidermal growth factor (EGF) levels from infants receiving breast milk, a rich source of the growth factor, were compared with the levels excreted by infants receiving bovine milk based formulae or total parenteral nutrition that contains very little EGF. Although at 5-10 days after birth there was no significant difference in the urinary EGF output by the three groups of infants, by 13-17 days the urinary EGF output by breastfed infants was higher than by infants fed the two EGF-poor diets. These latter results are consistent with the hypothesis that EGF crosses the gastrointestinal wall to enter the general circulation in the suckling infant.

Adolescent↗
PubMed1989

Sheep insulin-like growth factors I and II: sequences, activities and assays.

This report describes the purification, sequences, and activities of insulin-like growth factors (IGFs) from adult and fetal sheep plasma. IGF-1 from adult sheep is identical to human and bovine IGF-I, except for substitution in the sheep of Ala at residue 66 for Pro in the human and bovine polypeptides. IGF-II from adult sheep differs from bovine IGF-II also by a single amino acid, with residue 62 being Ala in ovine and Thr in bovine IGF-2. The first 10 amino-terminal residues of fetal sheep plasma IGF-I and 92% of the amino acids of fetal IGF-II were identified and found to be the same as those of the corresponding IGFs isolated from adult sheep. Ovine IGF-I was virtually equipotent with human IGF-I in growth-related bioassays and in a RIA for human and bovine IGF-I and inhibited the binding of radiolabeled human IGF-I to type I IGF receptors and to a pure IGF-binding protein. Ovine and bovine IGF-II were also found to be similar to each other in biological and immunochemical activities, and in their binding to type I and II IGF receptors and IGF-binding protein. As observed with human and bovine IGF-I and IGF-II, ovine IGF-I bound slightly better to type I IGF receptors than ovine IGF-II, but bound very poorly to type II IGF receptors. This study shows that IGFs from sheep are very similar to those of human and bovine in structure and activity and defines sensitive radioligand assays specific for ovine IGF-I and ovine IGF-II.

Amino Acid Sequence↗
PubMed1989

Purification, amino acid sequences and assay cross-reactivities of porcine insulin-like growth factor-I and -II.

Porcine insulin-like growth factor-I (IGF-I) and IGF-II have been characterized to help define the roles of these peptides in the growth process. The amino acid sequence of porcine IGF-I was found to be identical to the human and bovine peptides. Porcine IGF-II was more similar to human IGF-II than to forms of this growth factor in other mammalian species, differing only in the replacement of asparagine for serine at residue 36. In a biological assay that measures the stimulation of protein synthesis in rat L6 myoblasts, porcine IGF-I was approximately ninefold more potent than porcine IGF-II or bovine IGF-II, while recombinant human IGF-I and IGF-II had half the potency of the respective natural peptides. Porcine and recombinant human IGF-I showed essentially equal competition for binding in a human IGF-I radioimmunoassay while between 0.6 and 1.5% cross-reactivity was observed with human, bovine or porcine IGF-II. A receptor assay for IGF-II demonstrated similar potencies for the three IGF-II peptides, while the cross-reactivity of recombinant human IGF-I was only 0.05%. Porcine IGF-I exhibited a higher cross-reactivity, presumably due to very slight contamination with IGF-II.

Amino Acid Sequence↗
PubMed1989

Effects of insulin-like growth factors on protein metabolism: why are some molecular variants more potent?

The insulin-like growth factors (IGFs) produce a dual anabolic effect on protein metabolism in cultured cells via a stimulus of the synthetic pathway and an inhibition of the degradative pathway. Accordingly, they offer promise as agents that may retard the extensive and life-threatening negative nitrogen balance that accompanies human polytrauma. In this report we describe the discovery of a novel, more potent, form of IGF-1, des-(1-3)-IGF-1, and explain its increased action as resulting from higher concentrations of the free peptide produced because des-(1-3)-IGF-1 binds very poorly to IGF carrier proteins released from most cell types. The truncated growth factor retains a long biological half-life because it is attached to blood IGF-binding proteins. Further understanding of the biological significance of IGF, coupled with synthetic approaches directed at the production of mutant IGF-1 molecules, can be expected to yield significant advances in the treatment of polytrauma.

Animals↗
PubMed1988

Classification of the insulin-like growth factor binding proteins into three distinct categories according to their binding specificities.

Competitive binding experiments with insulin-like growth factor (IGF)-1, IGF-2 and des-(1-3)-IGF-1 have confirmed the interpretation based on limited amino-terminal sequence analysis that at least three types of IGF binding protein occur. In addition to the acid stable subunit of the large serum binding protein which exhibits des-(1-3)-IGF-1 binding only slightly less than IGF-1, the small IGF binding proteins can be separated into two classes based on differences in des-(1-3)-IGF-1 and IGF-2 binding potencies.

Animals↗
PubMed1988

Sequence and domain structure of yeast pyruvate carboxylase.

The nucleotide sequence of the yeast pyruvate carboxylase gene has been determined from a cloned fragment of yeast genomic DNA. The deduced translation product codes for a polypeptide of 1178 amino acids, having a calculated molecular weight of 130,100. The protein shows strong sequence homology to specific regions of other biotin carboxylases, lipoamide transferases, and carbamyl phosphate synthetases. The homologous regions suggest the presence of three subsites in the enzyme: a biotin attachment site, a keto acid-binding site, and an ATP-binding site. Partial proteolysis with a variety of proteases under nondenaturing conditions indicates the presence of structural domains corresponding to these subsites.

Adenosine Triphosphate↗
PubMed1988

Detection of protein similarities using nucleotide sequence databases.

A simple procedure is described for finding similarities between proteins using nucleotide sequence databases. The approach is illustrated by several examples of previously unknown correspondences with important biological implications: Drosophila elongation factor Tu is shown to be encoded by two genes that are differently expressed during development; a cluster of three Drosophila genes likely encode maltases; a flesh-fly fat body protein resembles the hypothesized Drosophila alcohol dehydrogenase ancestral protein; an unknown protein encoded at the multifunctional E. coli hisT locus resembles aspartate beta-semialdehyde dehydrogenase; and the E. coli tyrR protein is related to nitrogen regulatory proteins. These and other matches were discovered using a personal computer of the type available in most laboratories collecting DNA sequence data. As relatively few sequences were sampled to find these matches, it is likely that much of the existing data has not been adequately examined.

Alcohol Dehydrogenase↗
PubMed1988

Inactivation of chicken liver pyruvate carboxylase by 1,10-phenanthroline.

Inactivation of chicken liver pyruvate carboxylase by the chelating agent 1,10-phenanthroline follows pseudo-first-order kinetics. The hyperbolic dependence of the apparent first-order rate constant on 1,10-phenanthroline concentration is consistent with a two-step inactivation mechanism, in which 1,10-phenanthroline binds firstly to the enzyme, and secondly to the enzyme-bound Mn(II) ion. Binding of 1,10-phenanthroline to pyruvate carboxylase results in complete loss of ATP/Pi exchange activity, but only a 61% decrease in pyruvate/oxaloacetate exchange activity. The rate of inactivation is greater at low enzyme concentrations, implying that binding of 1,10-phenanthroline to monomers and dimers is preferred relative to that of tetramers. Furthermore, in the presence of acetyl-CoA, which stabilizes the tetrameric structure, no dependence of inactivation on enzyme concentration is observed. As monitored by gel-permeation liquid chromatography, formation of the enzyme-Mn(II)-phenanthroline complex results in loss of the tetrameric structure of the enzyme. From atomic-absorption measurements, inactivation by 1,10-phenanthroline also causes some loss of Mn(II) from the enzyme. It is concluded that the Mn(II) atom does not participate directly in the reaction mechanism, but may play a structural role essential to the integrity of the enzyme's tetrameric structure.

Acetyl Coenzyme A↗
PubMed1988

Ultraviolet irradiation of nucleic acids: formation, purification, and solution conformational analyses of oligothymidylates containing cis-syn photodimers.

Acetone-photosensitized UV irradiation of three thymine oligomers, d(TpT), d(TpTpT), and d(TpTpTpT), forms predominantly cis-syn cyclobutyl photodimers. C-18 reverse-phase high-performance liquid chromatography is used to purify the following positional isomers: d(TpT[p]T), d(T[p]TpT), d(TpTpT[p]T), d(TpT[p]TpT), d(T[p]TpTpT), and d(T[p]TpT[p]T), where T[p]T represents the cis-syn photodimer. Conformational properties of the cis-syn dimers and adjacent thymine nucleotides have been investigated in solution by using 1H, 13C, and 31P NMR spectroscopy. These studies show that (1) the photodimer conformation in longer oligothymidylates is similar to that in the dinucleoside monophosphate and (2) the cis-syn dimer induces alterations to a greater degree on the 5' side than on the 3' side of the photodimer. Specifically, the photodimer distorts the exocyclic bonds epsilon(C3'-O3') in Tp- and gamma(C5'-C4') in -pT[p]- on the 5' side and slightly alters the furanose equilibrium of the -pT nucleotide on the 3' side of the dimer.

Magnetic Resonance Spectroscopy↗
PubMed1988

Aggregation of lysine-containing zeins into protein bodies in Xenopus oocytes.

Zeins, the storage proteins of maize, are totally lacking in the essential amino acids lysine and tryptophan. Lysine codons and lysine- and tryptophan-encoding oligonucleotides were introduced at several positions into a 19-kilodalton zein complementary DNA by oligonucleotide-mediated mutagenesis. A 450-base pair open reading frame from a simian virus 40 (SV40) coat protein was also engineered into the zein coding region. Messenger RNAs for the modified zeins were synthesized in vitro with an SP6 RNA polymerase system and injected into Xenopus laevis oocytes. The modifications did not affect the translation, signal peptide cleavage, or stability of the zeins. The ability of the modified zeins to assemble into structures similar to maize protein bodies was assayed by two criteria: assembly into membrane-bound vesicles resistant to exogenously added protease, and ability to self-aggregate into dense structures. All of the modified zeins were membrane-bound; only the one containing a 17-kilodalton SV40 protein fragment was unable to aggregate. These findings suggest that it may be possible to create high-lysine corn by genetic engineering.

Amino Acid Sequence↗
PubMed1988

Insulin-like growth factors 1 and 2 in bovine colostrum. Sequences and biological activities compared with those of a potent truncated form.

1. Insulin-like growth factors 1 and 2 (IGF-1 and IGF-2) together with a truncated form of IGF-1 were purified to homogeneity from bovine colostrum. 2. Two forms of IGF-1 were totally resolved from IGF-2 in the purification by h.p.l.c. involving cation-exchange and reverse-phase columns. 3. The complete amino acid sequences for all three forms of IGF were determined. The sequence of bovine IGF-1 was found to be identical with that of human IGF-1, and that of the variant lacked the N-terminal tripeptide Gly-Pro-Glu (-3N:IGF-1). Bovine IGF-2 was found to differ in three residues of the C-domain compared with human IGF-2, with serine, isoleucine and asparagine substituted for alanine, valine and serine respectively at positions 32, 35 and 36. 4. Protein synthesis in L6 rat myoblasts was stimulated and protein degradation inhibited in a co-ordinate response with all three IGFs. The relative potency in both processes was -3N:IGF-1 greater than IGF-1 greater than IGF-2. A similar order of potency was obtained for the stimulation of DNA synthesis by -3N:IGF-1 and IGF-1. The approximately 10-fold effect on biological activity of removing the N-terminal tripeptide is unexpected in view of current information on IGF-1 structure and function.

Amino Acid Sequence↗
PubMed1988

The bovine insulin-like growth factor (IGF) binding protein purified from conditioned medium requires the N-terminal tripeptide in IGF-1 for binding.

The insulin-like growth factor binding protein (BP) secreted by bovine kidney (MDBK) cells has been purified by affinity chromatography on a rat IGF-2 Sepharose column. Purified BP migrated as a single band of Mr 40,000 upon SDS polyacrylamide gel electrophoresis. An N-terminal sequence of 53 residues was obtained which was very similar up to residue 21 to the corresponding rat BRL-3A BP sequence. In competitive binding experiments with bovine IGF-1 and IGF-2, and recombinant human IGF-1, BP had a similar affinity for these ligand when IGF-1 tracer was used, but a higher affinity for IGF-2 with IGF-2 as radioligand. The N-terminal destripeptide truncated form of bovine IGF-1, which has enhanced biological activity, was found to have a markedly reduced affinity for BP compared to intact IGF-1. The increased bioactivity of destripeptide IGF-1 can be explained by this reduced affinity for BP.

Animals↗
PubMed1988

A large family of bacterial activator proteins.

At least nine different bacterial proteins belong to the LysR family. The gene sequence for one of these proteins is presented here. Six others (Escherichia coli LysR, IlvY, CysB; Salmonella typhimurium MetR; Rhizobium NodD; and Enterobacter cloacae AmpR) are known to activate other genes. Based on sequence alignments, each member of this family is predicted to have a helix-turn-helix DNA binding motif near its amino terminus. The combined evidence indicates that all nine proteins are related by common ancestry, are similarly folded, and are not detectably related to other known bacterial regulatory proteins. The DNA database searching procedure and other methods used in this study should be useful in detecting other groups of related proteins.

Bacterial Proteins↗
PubMed1988

Radiation chemistry of d(TpApCpG) in oxygenated solution.

The radiation chemistry of the DNA tetranucleoside triphosphate d(TpApCpG) was investigated. The tetramer was X-irradiated in oxygenated aqueous solution and the various products separated by high-performance liquid chromatography. The principal modifications of the tetramer were analysed intact using nuclear magnetic resonance (NMR) spectroscopy and in some instances also by fast atom bombardment (FAB) mass spectrometry. The principal radiation-induced lesions of d(TpApCpG) were found to be a formamido modification derived from the thymine base, two stereoisomeric forms of a 1-carbamoyl-2-oxo-4,5-dihydroxyimidazolidine modification derived from the cytosine base and an 8-hydroxyguanine modification.

Chromatography, High Pressure Liquid↗
PubMed1988

Cerebral infarction pattern identified on emission computed tomography using technetium-99m MDP.

In the metastatic work-up of a patient with a malignant lung tumor and cerebrovascular disease, bone imaging was performed which showed abnormal uptake in the right parietal region. Emission computed tomography indicated that this was due to intracranial disease and revealed a wedge-shaped pattern indicative of cerebral infarction. A later follow-up study showed complete clearance of the abnormal uptake.

Cerebral Infarction↗