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T L Chang

Publications and source records attributed to T L Chang.

34 records · Page 2Linked to original sources
PubMed1983

Characterization of proopiocortin converting activity in rat anterior pituitary secretory granules.

Lysates from purified secretory granules of rat anterior pituitary glands were incubated with [3H]phenylalanine or [3H]arginine-labeled toad proopiocortin. The processed products formed were identified by immunoprecipitation with ACTH and beta-endorphin antisera, and by comigration with known markers on acid-urea polyacrylamide gels. Proopiocortin was cleaved by the secretory granule lysate primarily to 21,000 mol wt ACTH, 13,000 mol wt ACTH, 16,000 mol wt NH2-terminal glycopeptide, beta-lipotropin, a beta-endorphin-like peptide, and beta-endorphin. Characterization of the anterior pituitary proopicortin-converting activity shows that it: (1) cleaves specifically at the peptide bond on the carboxy side of the lysine-arginine residues of proopiocortin, (2) has a pH optimum in the acidic range, (3) is present in membrane and soluble fractions of the granule lysate, and (4) is inhibited by leupeptin, pepstatin A, and 2,2' dithiodipyridine, but not by p-chloromercuribenzoate, diisopropyl fluorophosphate, N alpha-p-tosyl-L-lysine chloromethyl ketone hydrochloride, chloroquine, L-1-tosylamide-2-phenylethyl-chloromethyl ketone, or EDTA.

Adrenocorticotropic Hormone↗
PubMed1982

Proopiocortin-converting enzyme activity in bovine neurosecretory granules.

Neurosecretory granules (NSGs) from neural lobes of bovine pituitary glands were isolated in a highly purified form by metrizamide-sucrose gradient centrifugation. The purified NSGs were lysed and centrifuged, and the supernatants were further fractionated by gel filtration on Sephadex G-75. Proopiocortin-converting enzyme activity was assayed by incubation of [3H]arginine- or [3H]phenylalanine-labeled toad proopiocortin with NSG supernatant fractions. The processed products were identified by immunoprecipitation with ACTH and beta-endorphin antisera, followed by acid-urea gel electrophoresis. The optimum pH for the enzyme-mediated conversion was around pH 5.0. Conversion of toad proopiocortin by NSG converting enzyme activity was inhibited by leupeptin, antipain, p-chloromercuribenzoate, and pepstatin A, but not by diisopropyl fluorophosphate, EDTA, or N-alpha-p-tosyl-L-lysine-chloromethyl ketone HCl. The results suggest that the proopiocortin-converting enzyme activity in bovine neurosecretory granules is due to an acid-thiol protease which may contain secondary hydrophobic binding sites that are involved in substrate recognition.

Adrenocorticotropic Hormone↗
PubMed1980

Stability differences between high coagulant (alpha) and noncoagulant (gamma) human thrombins. Denaturation.

Human alpha-thrombin, the two (covalently linked)-chain, highly coagulant blood-clotting enzyme was compared with its noncoagulant, yet estero/amidolytically active derivative, gamma-thrombin, a three (noncovalently associated)-domain enzyme which results from two proteolytic cleavages of the coagulant a form. Studies of their denaturation behavior by Tos-Arg-OMe esterase activity, by intrinsic fluorescence, by fluorescence of active serine-directed dansyl labels, and by monitoring the ESR of a fluorosulfonylphenyl spin-labeled inhibitor clearly demonstrated the reduced stability of the noncovalently associated gamma-thrombin form. At pH 6.5, 0.75 M NaCl, gamma-thrombin unfolds in approximately 2. 1 M urea while the more stable a form denatures at approximately 4 M urea. By monitoring active serine probes (spin label or fluorescent labels), these transitions were slightly lower, 1.0 +/- 0.1 and 2.8 +/- 0.2 M urea for spin-labeled gamma- and alpha-thrombins, respectively. Similar behavior was found for the same spin-labeled derivatives in guanidine HCl with unfolding transitions of 0.4 M and 1.0 M for spin-labeled gamma- and alpha-thrombin, respectively. These differences in structural stabilization serve as a good physical diagnostic for the two thrombin species.

Blood Coagulation↗
PubMed1980

Refolding of a three (noncovalently linked)-domain enzyme. Human gamma-thrombin.

Human gamma-thrombin is a three (noncovalently linked)-domain enzyme which contains the known serine protease catalytic triad, Asp-His-Ser, one on each of the three noncovalent domains (Asp 99 on the A-B3 chain). While protein-folding dogma does not necessarily predict that the denatured form of this enzyme could refold to the correct conformation, a monitor of the esterase activity (Tos-Arg-OMe) shows complete recovery of native catalytically active conformation. When compared with the covalently intact alpha form which refolds from urea in less than 2 min with complete return of both clotting and esterase activity, gamma-thrombin requires up to 90 min to regain full esterase activity. The gamma-thrombin reactivation data best fit a single first order rate constant, k = 0.03 +/- 0.005 min-1. It was suggested that the gamma-thrombin renaturation process might represent first the rapid refolding, then subsequent reassociation and reisomerization of the three noncovalent domains to yield a lower energy, fully active, conformation. This study represents the only example known of the refolding (reconstitution) of a three (noncovalent)-domain protein.

Blood Coagulation↗
PubMed1979

Cobalt iontophoresis of sensory nerves in the rat lung.

By iontophoretically introducing, first, cobalt and, subsequently, sulfide ions into the vagus nerve, it is possible to trace sensory nerves to their endings in the rat lung. Nerve fibers and terminals are found predominantly in the adventitia of the airways and blood vessels. Some nerves are found in the submucosa of the bronchi and bronchioles. Some are found in the cardiac muscle on the periphery of pulmonary veins, and a few nerves are seen to end among smooth muslces of the blood vessels and the airways. At least three types of nerve endings can be identified at the light microscopic level: (1) free nerve endings; (2) brush-like endings; (3) knob-like terminals.

Animals↗
PubMedDate not supplied

Pro-opiocortin processing in the pituitary: a model for neuropeptide biosynthesis.

The biosynthesis of alpha-MSH, beta-endorphin and ACTH in the pituitary is reviewed. These neuropeptides are synthesized from a common pro-protein, pro-opiomelanocortin. The pro-protein is cleaved intragranularly, at pairs of basic residues in the molecule to yield the respective peptide products. An unique, thiol protease (pro-opiocortin converting enzyme) and a carboxypeptidase B-like enzyme, both localized within pituitary secretory granules and having a pH optimum of 5-6, appear to be involved in the proteolytic processing of pro-opiomelanocortin.

Adrenocorticotropic Hormone↗