Search PubMed⌕ Search

Biomedical subjects

J T Dunn

Publications and source records attributed to J T Dunn.

At least 55 records · Page 3Linked to original sources
PubMed1989

Consensus sequences for early iodination and hormonogenesis in human thyroglobulin.

Thyroglobulin from a human goiter, containing four atoms of iodine/molecule (660,000 daltons), was iodinated with Na 125I and KI in vitro to achieve a net addition of either 2 or 7.8 atoms of iodine/molecule. After fractionation by high performance liquid chromatography, iodinated tryptic peptides from S-cyanoethylated 125I-thyroglobulin were purified, sequenced, characterized by [125I]iodoamino acid distribution, and localized within thyroglobulins primary structure based upon the published cDNA sequence, (Malthiery, Y., and Lissitsky, S. (1987) Eur. J. Biochem. 165, 491-498). The addition of 2 atoms of iodine/molecule of thyroglobulin produced iodotyrosyls at five principal sites, with no 125I-hormone formation. The addition of 7.8 atoms iodinated the same sites more heavily, produced iodotyrosyls at 10 additional sites, and formed iodothyronines at 5 sites. After addition of 2 atoms of iodine, tyrosyl 24 and 11% of thyroglobulins 125I, while tyrosyl 2572 had 24%, but with 7.8 added atoms of iodine, tyrosyl 24 had more of the thyroglobulins [125I]iodothyronine (36 versus 26%). Since tyrosyls 149, 866, and 1466 were iodinated early but did not form the inner rings of iodothyronines, they are attractive candidates for donors of outer iodothyronyl rings. The sequences around the iodotyrosyls fall into three consensus groups, as follows: 1) Glu/Asp-Tyr, associated with synthesis of thyroxine (residues 24, 2572, and 1309), or iodotyrosine (residues 2586 and 991); 2) Ser/Thr-Tyr-Ser, associated with synthesis of iodothyronine (residue 2765) and iodotyrosine (1466 and 883); and 3) Glu-X-Tyr, 7 of the remaining 8 iodotyrosyls occur in this sequence, and we found iodine incorporation at each place this sequence appears in the thyroglobulin molecule. Iodine has been found at homologues of most of these sites in thyroglobulins of other species. We conclude that the primary structure of thyroglobulin, and particularly these consensus sequences, have a major role in the formation of thyroid hormones and their iodinated precursors.

Amino Acid Sequence↗
PubMed1989

The hormonogenic sites of turtle thyroglobulin and their homology with those of mammals.

Thyroglobulin (Tg) from turtles previously injected with 125I was reduced, alkylated, and digested with trypsin. We purified the resultant peptides on HPLC columns, determined their amino acid sequences and the locations of [125I]T4 and [125I]T3 residues, and compared them with established sequences from humans, cows, rabbits, rats, and guinea pigs. We found five major T4 peptides, three of which were homologous with the major hormonogenic sites A, B, and D of mammalian Tg. Site A, the highly conserved major T4 site in mammals, had substitutions in three residues near the T4 residue and had much less of Tg's newly synthesized T4 than is found in mammalian Tg (25% in turtle vs. 44% in rabbit). Site B contained correspondingly more of Tg's new T4 (42% vs. 24% in rabbit). Turtle Tg contained little [125I]T3, and we did not find site C (Ser-T3/T4-Ser, the major T3 site in guinea pig and rabbit) in turtles, but did find Val-T4, a possible homolog. Site D was quantitatively less important than in mammals. The fifth turtle hormonogenic site, containing 12% of Tg's newly formed T4, had a tyrosyl residue substituted for the phenylalanine at residue 632 in the human sequence. We conclude that Tg's major hormonogenic sites are generally conserved across a considerable evolutionary distance, but that differences in primary structure occur and may contribute to changes in priority of hormone synthesis among these sites.

Alkylation↗
PubMed1988

Thyrotropin alters the utilization of thyroglobulin's hormonogenic sites.

We injected rabbits and guinea pigs with bovine thyrotropin (TSH) daily for 3 days, while controls received saline. All animals received sodium [125I]iodide on the second day, and thyroglobulin was purified from the thyroids of each group by gel filtration. Hormonogenic tryptic peptides from each S-cyanoethylated thyroglobulin preparation were isolated by high performance liquid chromatography, and their amino acid sequences were determined, permitting their localization within the thyroglobulin polypeptide chain by comparison with cDNA-derived sequences from bovine and human thyroglobulins. Thyroglobulins from the saline-injected rabbits and guinea pigs contained the same four major hormonogenic sites, designated A-D, previously described (Dunn, J. T., Anderson, P. C., Fox, J. W., Fassler, C. A., Dunn, A. D., Hite, L. A., and Moore, R. C. (1987) J. Biol. Chem. 262, 16948-16952). In both species, sites A and C were the major loci for thyroxine and triiodothyronine, respectively. However, site D in the guinea pig had a greater ratio of [125I]thyroxine to [127I]thyroxine than did site A, whereas the reverse was true in the rabbit. TSH administration produced the following changes in thyroglobulins of both species, relative to controls: 1) an increase in the ratio of [125I]triiodothyronine to [125I] thyroxine (rabbit, 0.29 versus 0.17; guinea pig, 0.19 versus 0.08), with the increase in triiodothyronine principally at site C; 2) a marked increase in 125I/127I and in thyroxine formation at site D (14.1% of thyroglobulin's thyroxine versus 9.8% in rabbits, 24 versus 13% in guinea pigs); 3) a corresponding decrease in thyroxine formation at site A (33 versus 43% in rabbits, 30 versus 46% in guinea pigs); and 4) a sharp increase in conversion of thyroglobulin's N-terminal 125I-labeled approximately 20 kDa hormone-rich iodopeptide, which contains site A, to a 125I-labeled approximately 15-kDa (rabbit) or 125I-labeled approximately 13-kDa (guinea pig) form, reflecting probable peptide bond cleavage. Our results show that TSH alters both the structure of the thyroglobulin molecule and the priority of utilization of its hormonogenic sites. We conclude that these changes are important to TSH's enhancement of thyroid hormone synthesis.

Amino Acid Sequence↗
PubMed1988

Cysteine proteinases from human thyroids and their actions on thyroglobulin.

This report describes properties of highly purified cathepsin-B and an additional cysteine proteinase, designated cysteine proteinase I, obtained from human thyroids. Both enzymes are localized to lysosomes. The activity profile of cysteine proteinase I combined with its sensitivity to the active site inhibitor Z-Phe-Phe-CNH2 suggest that it is distinct from other cysteine proteinases described so far. Cysteine proteinase I and cathepsin-B had respective pH optima of 3.5-4.0 and 4.5-5.0 with thyroglobulin (Tg) as substrate. Based on Km/Kcat (catalytic constant) ratios, cysteine proteinase I degraded rabbit [125I]Tg to peptide intermediates 50 times more efficiently than did cathepsin-B. Under conditions of limited digestion, both enzymes cleaved Tg at three or more sites, producing iodinated fragments of 20,000-50,000 mol wt (cysteine proteinase I) or 10,000-40,000 mol wt (cathepsin-B). Tryptic digests of these fragments were isolated by HPLC, and those containing thyroid hormone were sequenced for identification of amino acids and localization of 125I. Cysteine proteinase I cleaved peptides primarily from the C-terminal region of Tg, which contained two major hormonogenic sites, while cathepsin-B produced peptides mainly from the N-terminus, containing another major hormonogenic site. We suggest that the roles of cysteine proteinase I and cathepsin-B are the rapid initial fragmentation of Tg at opposite ends of the molecule, making hormone-containing sites accessible to additional cleavage by other lysosomal endopeptidases and exopeptidases.

Caseins↗
PubMed1988

Altered immunoreactivity of thyroglobulin in thyroid disease.

We prepared 3 samples of 19S thyroglobulin (Tg), 1 from a patient with Graves' disease, another from a patient with nontoxic goiter, and the third from a pool of Tg from normal subjects, and used each Tg preparation to produce a polyvalent antiserum in rabbits. The 3 antisera were similar to each other in their reactivity with thyroid Tg samples from 25 patients with various thyroid disorders and from 10 normal subjects. However, the immunoreactivity of the 35 individual Tg samples varied considerably. Decreased reactivity was associated with proteolysis during Tg preparation, iodination in vitro with 20 or more atoms of iodine/molecule Tg, the 27S species of Tg, Tg from 3 patients with thyroid cancer, and Tg from several patients with Graves' disease. The antiserum to Graves' Tg contained some antibodies that did not bind normal Tg on an affinity column, and these antibodies reacted more with Tg from patients with Graves' disease than with Tg from normal subjects or patients with nontoxic goiters. Thus, Tg from patients with Graves' disease may contain antigenic sites that are not present or exposed in Tgs from other subjects. We conclude that thyroid Tgs from patients with Graves' disease and from those with thyroid cancer may be different in structure from the Tgs of normal subjects. This conclusion is important to an understanding of Tg structure in thyroid disease and to the use of thyroid Tg for preparation of antisera and standards for measuring serum Tg concentrations.

Animals↗
PubMed1987

The sites of thyroid hormone formation in rabbit thyroglobulin.

Rabbit thyroglobulin (Tg) was labeled in vivo with 125I and purified by gel filtration. Separation by high performance liquid chromatography (HPLC) of tryptic digests of S-cyanoethylated Tg yielded four major iodothyronine-containing peaks, designated A, B, C, and D. These were further purified on HPLC and sequenced for identification of amino acid residues and for location of the iodothyronine by 125I counting. The published primary structure for bovine Tg, derived from cDNA sequencing of the Tg gene (Mercken, L., Simons, M.J., Swillens, S., Massaer, M., and Vassart, G. (1985) Nature 316, 647-651), permitted tentative location of the rabbit hormonogenic peptides within the Tg polypeptide chain. Site A, corresponding to bovine residue 5, contained 44% of Tgs [125I]T4 (thyroxine) and 25% of its [125I]T3 (triiodothyronine); its specific activity of iodine was higher than that for other sites, indicating priority of iodination. Site B, containing 24% of Tgs [125I]T4 and 18% of its [125I]T3, corresponded to bovine residue 2555. Site C, at the third residue from the C terminus (bovine residue 2748), was the major T3 site, accounting for over 50% of Tgs [125I]T3. The amino acid sequence around this site shows less homology among different animal species than do those flanking the other hormonogenic sites. Site D accounted for 17% of Tgs [125I]T4 and corresponded to bovine Tyr-1291, in the midportion of Tgs polypeptide chain. The three major T4-forming sites had the sequence Asp-Tyr (sites B and D) or Glu-Tyr (site A), while the sequence Ser-Tyr-Ser appeared to favor T3 synthesis (site C), suggesting an important influence of primary structure on hormonogenesis. We conclude that site A is the major T4-forming site and site C the major T3-forming one, but others are available and offer the opportunity for flexibility in meeting different demands for hormone formation.

Amino Acid Sequence↗
PubMed1986

Thyroid suppression and medical ablation for differentiated thyroid cancer.

Patients with thyroid cancer benefit from treatment with exogenous thyroid hormone for two reasons: it provides adequate levels of thyroid hormone to peripheral tissues, and it reduces the level of thyrotropin, which may be an important growth factor in patients with differentiated malignant neoplasms. The use of radioactive iodine for thyroid cancer is highly controversial. Its most appropriate applications are in follicular cancers, in older patients, and in distant functioning metastases. Its value in papillary cancer is questionable, particularly in young patients. There is a great need for effective basic and clinical research on the natural course of differentiated thyroid cancer and the effects of specific therapies.

Adenocarcinoma↗
PubMed1985

Management of injuries of the thoracic and abdominal aorta.

Thirty-five patients had surgery for injuries of the aorta at the Los Angeles County-USC Medical Center over a 4 1/2 year period. There were 27 survivors. The principles of management were to operate without delay if there was evidence of continued bleeding after initial fluid replacement as occurred in 11 patients. For the 24 patients who became stable after initial resuscitation, a more deliberate plan of management was used. Blood pressure was carefully monitored and controlled to avoid hypertension. Priorities for associated injuries were established and in several cases, they took treatment precedence over the aortic injury. Delay was sometimes necessary to utilize the more experienced personnel. In no instance did a stabilized patient hemorrhage during the delay. The most common injury seen was a blunt disruption of the proximal descending aorta. The details of the operative technique for this injury have been reported herein, along with a justification for not using either pump bypass or shunt to perfuse the distal aorta during the period of aortic cross-clamping.

Aorta, Abdominal↗
PubMed1984

The use of iodine as a thyroidal blocking agent in the event of a reactor accident. Report of the Environmental Hazards Committee of the American Thyroid Association.

In the event of a nuclear reactor accident, radioactive materials could be released into the environment: radioisotopes of iodine could constitute a major component of such a release. Upon such exposure, radioiodines could enter the body and accumulate in an unprotected thyroid gland where they would remain for varying periods of time. A number of methods have been proposed to protect those at risk of exposure. Administration of thyroid-blocking agents (such as potassium iodide) to exposed populations could be effective, but their use has raised a number of questions since there are considerable gaps in the scientific information available about the possible effects of low-level radiation from radioiodine. In addition, there are only limited data available about potential toxic side effects of potassium iodide distributed widely to large, unsupervised populations. Concern about these issues led the American Thyroid Association to appoint a committee of its members with special interest and competence in these areas to review the problems in detail and develop an advisory statement on the questions at issue for those to whom this matter might be of concern.

Accidents↗
PubMed1984

Roentgenographic contrast studies in acute small-bowel obstruction.

Plain abdominal roentgenograms are usually adequate for the treatment of patients with small-bowel obstruction. There is a select group of patients who may benefit from a more precise diagnosis or by nonoperative therapy for whom contrast roentgenograms may be helpful. Contrast roentgenograms were performed in 68 (21%) of 327 patients who had nondiagnostic abdominal films, atypical histories or findings, or those in whom nonoperative resolution would be highly desirable. Sixty-eight percent of 48 contrast tracers gave useful information, whereas 75% of 24 contrast enemas were helpful. Contrast material passed through a site of obstruction in 34 patients, yet 13 (38%) still required an operation. Contrast material did not pass in 16 patients and only one avoided a surgical procedure. No difference was noted between barium sulfate or water-soluble contrast material or how it was administered.

Adult↗
PubMed1984

Similar hormone-rich peptides from thyroglobulins of five vertebrate classes.

Thyroglobulins (Tgs) were purified from five species (rat, chicken, turtle, frog, and goldfish), each representing a different vertebrate class. On reduction with mercaptoethanol, each Tg produced five major iodopeptides, designated A-E, with ranges of estimated molecular mass, in kilodaltons (K), as follows: A, more than 300K; B, 210-280K; C, 30-42K; D, 19-28K; and E, 10-23K. Of these, the two smallest, D and E, had 40-80% of their iodine as iodothyronine, compared with 15-20% for the parent Tgs. They contained 25-63% of Tg's total iodothyronines but only a few percent of its peptide material. Calculations from amino acid analyses and iodine contents showed approximately 1 mol each of D and E/mol 660,000 dalton Tg. In comparisons of amino acid compositions by cluster analysis, iodopeptides D and E resembled each other and their counterparts in other species more than they resembled their parent Tgs. Also, the Tgs from different species were more similar to each other and to iodopeptides D and E than to nonthyroidal proteins randomly selected from the literature. We injected 125I into rats and turtles, and compared its distribution among the iodopeptides to that of 127I. These dual isotope experiments showed that as Tg was iodinated in vivo, iodopeptide B decreased both in molecular size and in its share of Tg's iodine, while the sum of iodopeptides D and E increased, indicating that B may be the precursor of D and E. In vivo iodination of rat Tg with 125I for different periods of time suggested that iodopeptide E and its iodothyronines are derived from a larger portion of the Tg molecule, perhaps iodopeptide B. The amount of 125I in iodopeptide D also increased with time. We conclude that these two discrete hormone-rich peptide fractions of small molecular weight are a consistent feature of Tg's structure throughout the vertebrate classes. Both are probably derived from a larger iodopeptide fraction (iodopeptide B) during the course of Tg maturation and iodination. Their conservation during evolution indicates an important role for them in thyroid hormone biosynthesis.

Animals↗
PubMed1984

Choice of therapy in young adults with hyperthyroidism of Graves' disease. A brief, case-directed poll of fifty-four thyroidologists.

We asked 54 thyroidologists how they would treat each of four patients having moderate hyperthyroidism of Graves' disease and a thyroid gland weighing 70 g (three to four times normal). For a 19-year-old woman, 67% of thyroidologists recommended an initial course of therapy with antithyroid drugs, usually for 1 year; 24% favored radioiodine treatments; and 9%, surgery. Choices for treating a 19-year-old man were similar. For a 29-year-old man, 44% of thyroidologists preferred drug therapy; 50%, radioiodine; and 6%, surgery. For a 29-year-old woman, choices were similar to those for the 29-year-old man, except for a slight preference for drugs over radioiodine. If hyperthyroidism recurred after a first course of antithyroid drugs, the consultants favored radioiodine treatments and surgery about equally, except in the 29-year-old man, in whom radioiodine was preferred. This survey shows considerable variation among experts in treating hyperthyroidism in young adults.

Adult↗
PubMed1983

The role of iodination in the formation of hormone-rich peptides from thyroglobulin.

Reduced thyroglobulins from several animal species contain hormone-rich iodopeptides of 20,000-26,000 and 15,000-18,000 daltons. The present study has investigated the role of iodination in their production. Experimental approaches have included: iodination in vitro of thyroglobulin from rabbit thyroid slices incubated with [3H]leucine and subsequent analysis of 3H distribution by gel electrophoresis; iodination in vitro of low iodine thyroglobulin from a human goiter, followed by isolation of the major iodopeptides and quantitation of their peptide content; and injection of iodine-deficient rats with Na125I and assessment of the distribution of isotope among the iodopeptides at successive time intervals. From these experiments the following general pattern has emerged: 1) at low levels of iodine (less than 5 atoms/molecule of thyroglobulin) in vitro or at short time intervals after iodine administration in vivo (less than 4 h), the principal iodinated component of reduced thyroglobulin is a approximately 230,000-dalton peptide; 2) with moderate increases in iodine (5-40 atoms/molecule) or longer time intervals after administration (greater than or equal to 4 h) there is less of the 230,000-dalton iodopeptide, and an iodothyronine-rich approximately 20,000- to 26,000-dalton iodopeptide appears; 3) at higher levels of iodine (greater than 40 atoms/molecule) the amount of approximately 230,000-dalton iodopeptide decreases further, the amount of 20,000- to 26,000-dalton iodopeptides may decrease, and an iodothyronine-rich 15,000- to 18,000-dalton peptide appears. Progressive iodination gives the same changes in distribution of peptide material among these iodopeptides as it does in iodine distribution, and the changes seen with iodination in vitro are similar to those occurring over time in vivo. We conclude that during the process of iodination discrete peptide bonds of thyroglobulin are cleaved to produce the hormone-rich iodopeptides. This is probably a normal part of thyroglobulin maturation in vivo and may be a necessary event preceding hormone formation.

Animals↗
PubMed1983

Mechanisms of activation of the classical pathway of complement by Hageman factor fragment.

The mechanism by which a fragment of activated Hageman factor (HFf) activates the classical pathway of complement in serum or platelet-poor plasma has been further delineated. When serum or platelet-poor plasma was incubated with various concentrations of HFf, the total complement hemolytic activity was reduced in a dose-dependent manner. This activation appears to be due to the direct interaction of HFf with macromolecular C1, since incubation of purified C1 with HFf resulted in dissociation of the subunits with concomitant reduction of C1r antigenicity that is indicative of C1 activation. HFf-dependent activation was prevented by prior treatment of HFf with the active site-directed inhibitor, H-D-proline-phenylalanine-arginine chloromethyl ketone or with a specific inhibitor of activated HF derived from corn. Incubation of HFf with highly purified C1r also resulted in activation of C1r as assessed directly using a synthetic substrate or indirectly by activation of C1s and consumption of C2. However, incubation of HFf with highly purified C1s resulted in formation of activated C1s (C1s-) but this was less efficient than HFf activation of C1r. We therefore conclude that activation of C1 in macromolecular C1 is the result of HFf conversion of C1r to C1r; activation of C1s then occurs primarily by C-1r and to a lesser degree by the direct action of HFf.

Amino Acid Chloromethyl Ketones↗
PubMed1982

The cleavage and formation of activated human Hageman factor by autodigestion and by kallikrein.

We have compared the cleavage of purified human Hageman factor (HF) by an activated form of human Hageman factor (HFa) (autodigestion) and by kallikrein. In each case, an initial cleavage is seen which produces HFa with Mr = 80,000 consisting of a heavy chain of Mr = 52,000 disulfide-linked to a light chain of Mr = 28,000. As autodigestion proceeds, HFa is shown to be further digested to yield a major active product at a molecular weight of 40,000 as well as Hageman factor fragment (HFf), which appear as two closely related molecular species of Mr = 28,000 and 30,000. A minor active product of Mr = 70,000 is also seen. Upon reduction of each of the active forms, a chain with Mr = 28,000 is released which contains the active site. HF digestion by kallikrein results in rapid formation of HFa, followed by HFa digestion to HFf and degradation of the heavy chain region to an inactive fragment at 40,000 daltons, which is then degraded to an end product of Mr = 36,000. Production of the active species with Mr = 40,000 and 70,000 is greatly diminished when kallikrein is the HF activator, and these active forms are shown to be formed primarily by autodigestion. The time course of HFa and HFf formation indicates that the rate of activation of Hageman factor by kallikrein is much faster than the rate of autoactivation; the addition of high molecular weight kininogen increases the rate of HFa and HFf formation as well as the extent of HG digestion. These data indicate that HFa is the active intermediate from which other active species are derived. The patterns of HF and HFa digestion by HFa and kallikrein are distinct; a model for HF digestion is presented.

Electrophoresis, Polyacrylamide Gel↗