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L C Huang

Publications and source records attributed to L C Huang.

54 records · Page 3Linked to original sources
PubMed1985

Phosphorylation of the regulatory subunit of type I cyclic AMP-dependent protein kinase by its catalytic subunit.

The regulatory subunit (R) of Type I cAMP-dependent protein kinase from rabbit skeletal muscle can serve as a substrate for its catalytic subunit (C). The degree of phosphorylation depends on both the concentration of C and the the time of incubation. Moreover, the phosphorylation can be totally blocked by protein kinase inhibitors. In contrast, cAMP stimulates the phosphorylation of R using the holoenzyme. The purified holoenzyme isolated from rabbit skeletal muscle can be further fractionated into two fractions on DEAE Sephadex column. The first fraction eluted with low salt (50 mM NaCl) contains a much lower concentration of kinases than the second fraction eluted with high salt (100 mM NaCl), but the low salt kinase can be readily phosphorylated in the presence of MgATP. Our data thus implies that only a small fraction of Type I cAMP-dependent protein kinases in the skeletal muscle is present as the phosphorylatable species.

Animals↗
PubMed1983

Chemical cross-linking of cyclic AMP-dependent protein kinase and its dissimilar subunits.

Previous kinetic studies have demonstrated that the activation of cyclic AMP-dependent protein kinase by cyclic AMP involves the formation of a ternary complex of cyclic AMP, the regulatory subunit (R) and the catalytic subunit (C). It is suggested that only this ternary complex breaks down to liberate the enzymically active catalytic subunit. We have performed cross-linking experiments with the holoenzyme and its dissimilar subunits in the presence of MgATP and various concentrations of cyclic AMP. Results from these cross-linking studies indicate that regulatory subunits exist as dimers in the native form. Moreover, dissociation of the holoenzyme or the reconstituted enzyme is promoted by cyclic AMP, and the effect of MgATP is to stabilize the enzyme in the tetrameric form. The success in cross-linking the regulatory and the catalytic subunits of protein kinase with the lysine-specific bifunctional cross-linking reagent dimethyl suberimidate may be attributed to the presence of a large number of lysine residues in the enzyme.

Adenosine Triphosphate↗
PubMed1983

Monoclonal antibody to human eosinophils recognizing 95 kD surface membrane antigen.

Mice were immunized with purified eosinophils obtained from patients with the idiopathic hypereosinophilic syndrome. A hybridoma initially producing an IgM antibody which switched to an IgG1 antibody was selected for cloning and further testing. This IgG1 antibody reacted with human eosinophils, granulocytes, monocytes and large granular lymphocytes, but did not react with T lymphocytes, B lymphocytes, platelets, erythrocytes, or a panel of human leukemia cells and cell lines. Bone marrow analysis revealed staining of myeloid precursor cells but not erythroid precursors or plasma cells. This IgG1 antibody had no effect on aggregation of granulocytes, lysozyme release, superoxide production, chemotaxis, or killing activity; however, there was some stimulation of beta-glucuronidase secretion. While the antibody did not augment the killing of Staphylococcus aureus by granulocytes, the antibody itself was bactericidal. By immunoprecipitation of granulocytes, eosinophils and monocytes, a molecule with a molecular weight of 95 kD was identified.

Animals↗
PubMed1980

Modification and identification of glutamate residues at the arginine-recognition site in the catalytic subunit of adenosine 3' :5'-cyclic monophosphate-dependent protein kinase of rabbit skeletal muscle.

It has been proposed that the active centre of cyclic AMP-dependent protein kinase contains an arginine-recognition site, which is considered to be essential for the function of the catalytic subunit of the kinase [Matsuo, Huang & Huang (1978) Biochem. J.173, 441-447]. The catalytic subunit can be inactivated by 3-(3-dimethylaminopropyl)-1-ethylcarbodi-imide and glycine ethyl ester at pH6.5. The enzyme can be protected from inactivation by preincubation with histone, a protein substrate of the enzyme. On the other hand, ATP, which also serves as a protein kinase substrate, does not afford protection. Polyarginine, a competitive inhibitor of protein kinase, which is known from kinetic studies to interact specifically with the arginine-recognition site, partially protects the catalytic subunit from inactivation by 3-(3-dimethylaminopropyl)-1-ethylcarbodi-imide. These results lead to the conclusion that the site of modification by carbodi-imide/glycine ethyl ester is most likely located at the arginine-recognition site of the active centre. A value of 1.7+/-0.2 (mean+/-s.d.) mol of carboxy groups per mol of catalytic subunit has been obtained for the number of essential carboxy groups for the function of protein kinase; a complete chemical modification of these essential carboxy groups results in total loss of catalytic activity. Finally, we have identified the essential carboxy group in the catalytic subunit of cyclic AMP-dependent protein kinase as being derived from glutamate residues. This is achieved by a three-step procedure involving an extensive proteolytic digestion of the [1-(14)C]glycine ethyl ester-modified enzyme and two successive high-voltage electrophoreses of the hydrolysate. It is concluded that 1.7mol of glutamyl carboxy groups per mol of catalytic subunit may be considered a component of the arginine-recognition site in the active centre of cyclic AMP-dependent protein kinase.

Amino Acids↗
PubMed1980

Kinetic studies on the dissociation of adenosine cyclic 3',5'-monophosphate from the regulatory subunit of protein kinase from rabbit skeletal muscle.

The exchange rate of unlabeled adenosine 3',5'-monophosphate (cAMP) with labeled [3H]cAMP in the dimeric regulatory subunit-cAMP complex of cAMP-dependent protein kinase, type I, purified from rabbit skeletal muscle is described by using the equilibrium isotope exchange technique. Results indicate that the rate of exchange carried out in the absence of the catalytic subunit (C) is rather slow with a half-life of approximately 870 s. This slow exchange rate is not affected by the presence of MgATP (50 microM). However, when both MgATP (50 microM) and C (1-13 NM) are present, the rate of isotope exchange is observed to increase markedly. Furthermore, less than stoichiometric amounts of C are required for the increase in the rate of cAMP exchange, indicating that the effect of C on the rate enhancement is a catalytic process. These results indicate that in the presence of MgATP, a ternary complex between C and regulatory subunit-cAMP complex must be formed, and a dynamic equilibrium between the eternary complex and its dissociable species must be reached within seconds. On the basis of our kinetic data, it is proposed that the formation of this ternary complex intermediate allows the rapid activation or the inactivation of cAMP-dependent protein kinase following changes in the cellular cAMP levels.

Animals↗
PubMed1980

Stimulation of muscle glycogen synthase phosphatase by polyamines.

The naturally-occurring polyamines were found to stimulate glycogen synthase phosphatase from rat skeletal muscles. The sequence of effectiveness in the stimulation was spermine greater than spermidine greater than putrescine. It was shown that the spermine-sensitive phosphatase was present primarily in the soluble fraction of the muscle extract. In the presence of spermine, the phosphatase activity can be further stimulated by Mn2+; however, a lower Mn2+ concentration is required for the activation of the enzyme in comparison with that in the absence of spermine. Kinetic studies indicated that activation of glycogen synthase phosphatase by spermine was achieved by an increase in its V without significant alteration in the Km, suggesting that spermine directly stimulated the catalytic efficiencly of the phosphatase enzyme.

Animals↗
PubMed1980

Effects of insulin treatment on the activities of phosphoprotein phosphatase and its inhibitors.

An increase in glycogen synthase phosphatase (phosphoprotein phosphatase) activity was observed in the rat skeletal muscle extract following insulin administration. The phosphoprotein phosphatase activity present in the muscle extract from insulin treated rats was observed to remain elevated after the extract had been subjected to a molecular sieve chromatography. These results indicate that the stimulatory effects of insulin is due to modification of phosphatase itself or some macromolecular weight modifiers. The heat-stable protein inhibitors of the phosphoprotein phosphatase were isolated from skeletal muscle of insulin treated and control rats and their inhibitory potencies were compared over a wide range of protein concentrations. The inhibitory potency in the insulin treated rat skeletal muscle was found to be significantly less than that in the control muscle. Since type-1 inhibitor is well-known to be active only after being phosphorylated by cyclic AMP-dependent protein kinase, we suggest that the observed change in phosphoprotein phosphatase inhibitor potency is most likely mediated by an alteration in the phosphorylation state of type-1 inhibitor.

Animals↗
PubMed1979

Hormonal control of glycogen metabolism.

In summary, we have presented evidence which relates to the action pathway of hormonal control of glycogen metabolism. In the case of insulin, there are changes demonstrable in the cyclic AMP-dependent protein kinase and also in the phosphoprotein phosphatase, under conditions where no direct relationship to either cyclic AMP or cyclic GMP levels are measurable. Therefore, a new unknown intermediate or second messenger system is again proposed. An insulin-generated labile compound(s) which inhibits the protein kinase has been discovered. This may function as an intermediate. Finally, the fact that the glycogen synthase system clearly differs from phosphorylase in its regulation by covalent phosphorylation is discussed. Synthase is now accepted as a multiply phosphorylated subunit, in contrast to phosphorylase which is singly phosphorylated. The inherent theoretical advantages of multiple phosphorylation over single phosphorylation are considered. The advantages of a multistate over a two-state model of enzyme interconversion are mentioned. The importance of the multiple phosphorylations interacting in a nonlinear manner with the control by cellular metabolites is in the explanation of how a small change in covalent phosphorylation signalled by a hormone can be translated in the cell milieu into a much larger change in rate.

Animals↗
PubMed1978

Evidence for an essential arginine recognition site on adenosine 3':5'-cyclic monophosphate-dependent protein kinase of rabbit skeletal muscle.

On the basis of the chemical and structural features of the amino acid sequences in the vicinities of phosphorylatable hydroxyamino acid residues in several of the well-known protein substrates for skeletal-muscle cyclic AMP-dependent protein kinase, it is hypothesized that the phosphorylatable residue at position i and arginine residue at position i-3 of these protein substrates are located on a peptide turn on the hydrophilic protein surface. It is further hypothesized that there is an arginine-recognition site near the active centre on the protein kinase. This site is essential for the function of cyclic AMP-dependent protein kinase, for, not only does it recognize specifically the exposed arginine residue of the protein substrate, but, more importantly, via the interaction with arginine-(i--3), it may help to steer the topologically adjacent serine-i into proper orientation on the nearby active centre for phosphorylation. Model-building and kinetic data that provide support for the proposed hypotheses are presented.

Adenosine Triphosphate↗
PubMed1975

Rabbit skeletal muscle protein kinase. Conversion from cAMP dependent to independent form by chemical perturbations.

Protein kinase isolated from rabbit skeletal muscle can be reversibly converted from the cAMP dependent form to the indepent form by chaotropic salts and urea. A similar but irreversible conversion can also be induced by trypsin digestion of the holoenzyme. The dissociation of cAMP dependent protein kinase by low concentrations of thiocyanate raises the possibility of isolating both native regulatory and catalytic subunits. From various changes in enzymatic activity caused by urea and trypsin perturbation, it is proposed that the conversion of protein kinase from the cAMP dependent to the independent form is due primarily to preferential modification of the regulatory subunit of the holoenzyme.

Animals↗
PubMedDate not supplied

Ethnic and geographic variations in the prevalence of hepatitis A, B and C among aboriginal villages in Hualien, Taiwan.

BACKGROUND: Taiwan is endemic for viral hepatitis infections. A field survey was performed in the isolated aborigines in Hualien, eastern Taiwan, to investigate the geographic and ethnic variations in hepatitis epidemiology. MATERIALS AND METHODS: From 1996 to 1998, blood was drawn from 1,748 subjects from two southern Ami and two northern Atayal villages for serum markers of hepatitis A, B and C. RESULTS: Hepatitis A infection approached 100% in all groups. Hepatitis B infection and carrier rates were higher in the Atayal than in the Ami (92.3% vs. 49.1% and 20.8% vs. 5.3%; p<0.01). Hepatitis C infection rates were higher in three villages (27.5%, 20.1% and 25.4% vs. 3.6%; p<0.01). Hepatitis C infection increased with age (p<0.01) while hepatitis B infection did not. CONCLUSION: Hepatitis A infected most aborigines before the age of 15 years. Hepatitis B seldom infected people after the age of 15 years, while hepatitis C continued to infect people who were older. Geographic factors are important for hepatitis C infection, whereas for hepatitis B infection, in addition ethnicity is also important.

Adolescent↗
PubMedDate not supplied

Primary ovarian cancer at National Taiwan University Hospital: 1980-1989.

BACKGROUND: Ovarian cancer is a silent killer. Most of the patients came with advanced stages. The retrospective review here was for a better understanding of the current situation of ovarian cancer patients treated at National Taiwan University Hospital. METHODS: One hundred and eighty-seven patients with primary ovarian cancer were treated at National Taiwan University Hospital from 1980 to 1989. Medical records were reviewed thoroughly. The WHO histological classification and FIGO staging system (1987) were used. RESULTS: The ages at diagnosis ranged from 2 to 87 years. Peak incidence was noted between 50-59 years. According to the WHO histological classification, the distribution of histological types included 79.1% epithelial, 4.3% sex-cord stromal tumors, 0.5% lipid cell, 12.8% germ cell, and 3.2% non-specific soft tissue origin. Based on FIGO staging system, 31.6% of the patients were in stage I, 11.2% in stage II, 40.6% in stage III and 16.6% in stage IV. Mean ages for common epithelial, non-epithelial, and germ cell ovarian cancers were 49, 26, 23 years, respectively. Cytoreductive surgery was done in 47.4% of stage I, 23.8% of stage II, 15.8% of stage III, and 12.9% of stage IV. The leading treatment modality was surgery plus chemotherapy, which was performed in 50.3% of whole series. Survival in patients with ovarian cancer is a function of clinical stage and histologic type. The 5-year observed survival rates for stage I to IV were 76.3%, 59.9%, 9.1%, and 4.3% respectively. The 5-year survival rate was 29.4% for epithelial ovarian cancer, 51.4% for non-epithelial ovarian cancer, and 64.5% for germ cell tumors. CONCLUSIONS: The prognosis and survival rate are still unsatisfactory despite the various modalities of treatment. Further improvements need the development of methods for early detection and multiple modalities of therapy.

Adolescent↗