scholarly journals The regulatory proteins of the myofibril. Characterization and properties of the inhibitory factor (troponin B)

1971 ◽  
Vol 123 (3) ◽  
pp. 367-377 ◽  
Author(s):  
M. C. Schaub ◽  
S. V. Perry
Keyword(s):  
Molecular Weight ◽  
Biological Activity ◽  
Inhibitory Activity ◽  
Gel Filtration ◽  
Inhibitory Factor ◽  
Regulatory Proteins ◽  
Actomyosin Atpase ◽  
Troponin Complex

1. Gel-filtration results indicate that the major component of inhibitory-factor preparations isolated by dissociation of the troponin complex consisted of a protein of subunit weight 23000 daltons. By the same procedure a molecular weight of 18000 was obtained for the calcium-sensitizing factor. 2. The inhibitory factor is specific for the actomyosin type of ATPase and ITPase. It is effective on desensitized actomyosin, natural actomyosin and intact myofibrils. 3. For inhibition, the actomyosin ATPase must be stimulated by Mg2+, Ca2+ or Mn2+. The Co2+-, Cd2+- or Zn2+-stimulated ATPases are not affected. 4. Biological activity is stable to treatment with dissociating agents, heat, pH11, pH1 and carboxymethylation. 5. Increasing amounts of actin, but not myosin or tropomyosin, progressively neutralize the inhibitory activity when added to desensitized actomyosin or myofibrils.

scholarly journals The regulatory proteins of the myofibril. Characterization and biological activity of the calcium-sensitizing factor (troponin A)

1972 ◽  
Vol 126 (1) ◽  
pp. 237-249 ◽  
Author(s):  
M. C. Schaub ◽  
S. V. Perry ◽  
W. Häcker
Keyword(s):  
Amino Acids ◽  
Biological Activity ◽  
Electrophoretic Mobility ◽  
Prolonged Exposure ◽  
Gel Filtration ◽  
Myofibrillar Protein ◽  
Inhibitory Factor ◽  
Regulatory Proteins ◽  
Protein System ◽  
Troponin Complex

1. Electrophoretically homogeneous calcium-sensitizing factor was prepared from the troponin complex by chromatography successively on sulphoethyl-Sephadex and on diethyl-(2-hydroxypropyl)aminoethyl-Sephadex in 6m-urea. It is a protein containing 53% of polar amino acids, of which a net excess consists of acidic residues. 2. On gel filtration the calcium-sensitizing factor was shown to be the only myofibrillar protein that bound 45Ca2+ tightly in the presence of 2–6m-urea. 3. Calcium-sensitizing factor effectively neutralized the effect of the inhibitory factor on the ATPase activities of actomyosin systems. Tropomyosin was essential for the regulation, by changes in the Ca2+ concentration, of the neutralizing effect of calcium-sensitizing factor on the inhibitory factor. 4. Prolonged exposure to chelators of Ca2+ produced an irreversibly modified form of calcium-sensitizing factor of higher electrophoretic mobility at pH8.6. The modified form neutralized the inhibitory factor action but this property could no longer be controlled by the Ca2+ concentration in the presence of tropomysin. 5. The calcium-sensitizing factor and tropomyosin could be replaced by their carboxymethylated derivatives in the relaxing-protein system.

scholarly journals The regulatory proteins of the myofibril. Separation and biological activity of the components of inhibitory-factor preparations

1972 ◽  
Vol 127 (1) ◽  
pp. 215-228 ◽  
Author(s):  
J. M. Wilkinson ◽  
S. V. Perry ◽  
H. A. Cole ◽  
I. P. Trayer
Keyword(s):  
Molecular Weight ◽  
Active Component ◽  
Inhibitory Factor ◽  
Regulatory Proteins ◽  
Additional Component ◽  
Limited Evidence ◽  
Molecular Weights ◽  
Molar Basis ◽  
Troponin Complex ◽  
Cysteine Content

1. Inhibitory-factor preparations isolated from myofibrils were shown to consist principally of proteins with molecular weights of 37000 and 23000. Under certain preparative procedures an additional component of molecular weight 14000 was present. 2. The 23000-dalton protein, the inhibitory factor, was the major active component. Its activity was enhanced by tropomyosin. 3. The 14000-dalton component also possessed inhibitory activity, although less than that of the 23000-dalton component when compared on a molar basis. Its activity was not always enhanced by tropomyosin. The 14000-dalton component could not be detected in whole fresh myofibrils and the limited evidence available is compatible with its formation during the preparation of the troponin complex. 4. The 37000-dalton component could not replace the inhibitory factor, calcium-sensitizing factor or tropomyosin as components of the relaxing-protein system. 5. All three components had distinctive amino acid compositions, particularly in their cysteine content.

A Potent Insect Chitinase Inhibitor of Fungal Origin

2003 ◽  
Vol 58 (11-12) ◽  
pp. 891-894 ◽  
Author(s):  
Teruhiko Nitoda ◽  
Hirokazu Usuki ◽  
Hiroshi Kanzaki
Keyword(s):  
Molecular Weight ◽  
Anion Exchange ◽  
Culture Filtrate ◽  
Inhibitory Activity ◽  
Spodoptera Litura ◽  
Gel Filtration ◽  
Fungal Strain ◽  
Water Soluble ◽  
Gel Filtration Chromatography ◽  
Soluble Polysaccharide

Abstract A water-soluble polysaccharide was isolated from the culture filtrate of a fungal strain, Sphaeropsis sp. TNPT116-Cz, as a novel insect chitinase inhibitor. It was purified to chromatographic homogeneity by ethanol precipitation, anion-exchange and gel filtration chromatography. Its molecular weight was estimated to be 16 kDa by gel filtration HPLC. Monosaccharide analysis showed that it contained glucose, galactose, N-acetylglucosamine and a deoxysugar. This polysaccharide showed potent and specific inhibitory activity against Spodoptera litura chitinase with an IC50 value of 28 nᴍ.

scholarly journals Circular-dichroic studies on the conformational behaviour of troponin and tropomyosin from bovine cardiac muscle

1978 ◽  
Vol 175 (1) ◽  
pp. 137-147 ◽  
Author(s):  
T I Lin ◽  
J Y Cassim
Keyword(s):  
Molecular Weight ◽  
Biological Activity ◽  
Amino Acid ◽  
Conformational Changes ◽  
Cardiac Troponin ◽  
Surface Charges ◽  
Troponin Complex ◽  
Induced Changes ◽  
Circular Dichroic

Bovine cardiac troponin is similar to rabbit skeletal troponin with respect to secondary structure, amino acid composition and molecular weight of the subunits, but differs slightly with respect to biological activity and surface charges of the subunits. Previous circular-dichroic studies of the subunits and recombination of subunits have indicated significant Ca2+-induced delocalized conformational changes. Present studies of the native troponin complex are not in accord with such changes. Furthermore the formation of the troponin-tropomyosin complex in vitro results in no delocalized conformational changes, nor does it sensitize troponin to Ca2+-induced changes. It is suggested that the troponin complex cannot be dissociated into subunits without significant and irreversible conformational perturbation.

Inhibitors of Urokinase-Induced Fibrinolysis in Pregnancy

1979 ◽  
Author(s):  
J. E. Duthie ◽  
D. M. Campbell ◽  
D. Ogston
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Inhibitory Activity ◽  
Substantial Reduction ◽  
Gel Filtration ◽  
Late Pregnancy ◽  
Lower Molecular Weight ◽  
Α2 Macroglobulin ◽  
Molecular Weight Fractions ◽  
In Pregnancy

Pregnancy plasma possesses inhibitory activity against urokinase measured on unheated fibrin plates. Antiurokinase activity in late pregnancy plasma subjected to gel filtration on Sephadex G-200 eluted with the high molecular weight proteins including α2-macroglobulin and, in greater quantity, with albumin. In all non-pregnancy plasmas the high molecular weight inhibitor activity was present in equivalent quantities; the lower molecular weight inhibitor was found in small amounts in only a proportion of plasmas. The anti-urokinase activity of pregnancy plasma could be separated from α1-antitrypsin and α2-antiplasmin by chromatography on DEAE-Sephadex. Within 1 hour of parturition plasma fibrinolytic activity increased and there was substantial reduction in the anti-urokinase activity of the lower molecular weight fractions; no change was seen in the high molecular weight inhibitory activity. It is concluded that anti-urokinase activity in pregnancy plasma resides in a protein distinct from established protease inhibitors; a placental source is postulated.

Inhibition of tryptophan synthase by extracts from pea roots

1969 ◽  
Vol 47 (5) ◽  
pp. 649-653 ◽  
Author(s):  
James Chen ◽  
W. G. Boll
Keyword(s):  
Molecular Weight ◽  
Gel Filtration ◽  
Inhibitory Factor ◽  
Heat Inactivation ◽  
Tryptophan Synthase ◽  
Fresh Tissue ◽  
Low Concentration ◽  
Root Extracts ◽  
Pea Roots ◽  
Inhibitory Components

Low tryptophan synthase (L-serine hydro-lyase (adding indole), EC 4.2.1.20) activity in extracts of pea roots is a consequence of both a low concentration of the enzyme in roots and the presence of inhibitors at least some of which may be removed by homogenizing the fresh tissue with acetone. Inhibition by root extracts, and fractions thereof, was assayed against tryptophan synthase from seedling buds. At least three inhibitory components were detected, namely (1) a dialyzable, thermostabile component, (2) an undialyzable, thermolabile component, and (3) an undialyzable, thermostabile component. A dialyzable factor which protects an undialyzable inhibitory factor against heat inactivation is also indicated.Fractionation of crude root extracts by gel filtration on a column of Sephadex G-50 shows inhibitor with a molecular weight of about 10 000 or larger.

scholarly journals The contractile and regulatory proteins of insect flight muscle

1973 ◽  
Vol 135 (2) ◽  
pp. 277-286 ◽  
Author(s):  
Belinda Bullard ◽  
Renata Dabrowska ◽  
Lowell Winkelman
Keyword(s):  
Molecular Weight ◽  
Atpase Activity ◽  
Flight Muscle ◽  
Insect Flight ◽  
Ph Dependence ◽  
Regulatory Protein ◽  
Gel Filtration ◽  
Regulatory Proteins ◽  
Insect Flight Muscle ◽  
Subunit Molecular Weight

1. Myosin, actin and the regulatory proteins were prepared from insect flight muscle. 2. The light subunit composition of the myosin differed from that of vertebrate muscle myosin. The ionic strength and pH dependence of the myosin adenosine triphosphatase (ATPase) were measured. 3. Actin was associated with a protein of subunit molecular weight 55000 and was purified by gel filtration. Impure actin had protein bound at a periodicity of about 40nm. 4. Regulatory protein extracts had tropomyosin and troponin components of subunit molecular weight 18000, 27000 and 30000. Crude extracts of regulatory proteins inhibited the ATPase activity of desensitized or synthetic actomyosin; this inhibition was relatively insensitive to high Ca2+ concentrations. Purified insect regulatory protein produced as much sensitivity to Ca2+ as did the rabbit troponin–tropomyosin complex. 5. Synthetic actomyosins were made from rabbit and insect proteins. Actomyosins containing insect myosin had a low ATPase activity that was activated by tropomyosin. The Ca2+ sensitivity of actomyosins containing insect myosin or actin, with added troponin–tropomyosin complex from rabbit, was comparable with that of rabbit actomyosin.

scholarly journals Biosynthesis of glucagon in isolated pancreatic islets of guinea pigs

1974 ◽  
Vol 140 (1) ◽  
pp. 13-21 ◽  
Author(s):  
Claes Hellerström ◽  
Simon L. Howell ◽  
John C. Edwards ◽  
Arne Andersson ◽  
Claes-Göran Östenson
Keyword(s):  
Molecular Weight ◽  
Biological Activity ◽  
Gel Filtration ◽  
Subcellular Fractionation ◽  
Fat Cell ◽  
Electron Microscopic ◽  
Isolated Pancreatic Islets ◽  
Isolated Islets Of Langerhans ◽  
Electron Microscopic Radioautography ◽  
Very High

1. The biosynthesis of glucagon in guinea-pig A2 cells was investigated by incubation of isolated islets of Langerhans in the presence of [3H]tryptophan for periods of up to 14 days. Proteins were extracted from islets and incubation media and analysed by gel filtration. 2. In addition to very-high-molecular-weight (100000) proteins, the principal tryptophan-containing biosynthetic product after incubation for up to 17h was a protein of minimum mol.wt. 9000, which co-eluted on gel filtration with a peak of glucagon-like immunoreactivity, but was apparently devoid of biological activity in a fat-cell assay. A discrete peak of labelled glucagon was only recovered after incubation for at least 6 days. Losses of glucagon during the extraction and rapid secretion of newly synthesized glucagon into incubation media were excluded as reasons for the lack of recovery of labelled hormone from islets after shorter incubations. 3. The 9000-mol.wt. protein was localized to A2 cells in experiments using B-cell-depleted islets, and to A2-cell granules by subcellular fractionation and electron-microscopic radioautography. Only glucagon was secreted into the incubation medium. 4. Possible relationships between the 9000-mol.wt. protein and glucagon are discussed in the light of postulated mechanisms of glucagon biosynthesis.

scholarly journals STUDIES ON THE PATHOGENESIS OF FEVER

1968 ◽  
Vol 127 (2) ◽  
pp. 341-357 ◽  
Author(s):  
Marilyn Sue Kozak ◽  
Helmut H. Hahn ◽  
William J. Lennarz ◽  
W. Barry Wood
Keyword(s):  
Molecular Weight ◽  
Biological Activity ◽  
Active Material ◽  
Gel Filtration ◽  
Chemical Properties ◽  
Disc Electrophoresis ◽  
Polyacrylamide Gel ◽  
Sulfhydryl Groups ◽  
Biologically Active ◽  
Exhaustive Extraction

Small quantities of highly purified granulocytic pyrogen have been separated from contaminating proteins by disc electrophoresis in polyacrylamide gel. The biologically active material thus isolated was shown to be electrophoretically homogeneous at pH 9 and pH 3.8. Earlier work on the chemical properties of the pyrogen molecule has been extended to include: (a) estimation of its molecular weight by gel filtration; (b) demonstration of free sulfhydryl groups essential for its biological activity; and (c) evidence that it is not inactivated by exhaustive extraction with ethanolether or n-heptane.

DISCREPANCIES IN PLASMA LH ACTIVITIES AS MEASURED BY RADIOIMMUNOASSAY AND AN IN VITRO BIOASSAY

1974 ◽  
Vol 77 (4) ◽  
pp. 672-685 ◽  
Author(s):  
M. H. Qazi ◽  
P. Romaní ◽  
E. Diczfalusy
Keyword(s):  
Molecular Weight ◽  
Biological Activity ◽  
Gel Filtration ◽  
Biologically Active ◽  
Plasma Samples ◽  
Elution Volume ◽  
Immunological Activity ◽  
Molecular Weight Range ◽  
Weight Range

ABSTRACT Using a highly sensitive in vitro bioassay system, luteinizing hormone activity has been measured in parallel with radioimmunoassays in postmenopausal plasma and plasma obtained from women in various phases of the menstrual cycle (follicular, mid-cycle, luteal). Biological and immunological activities were measured directly in plasma samples without any chemical manipulation. The biological activity (B) was always higher than the immunological activity (I); the B/I ratio varied from 2.1 to 14.0. Gel filtration of pooled plasma samples through Sephadex G-100 revealed major discrepancies in each physiological state when immunological and biological activities were measured in each fraction. The biological activity was eluted as a single peak behind the elution volume of bovine serum albumin, but in front of the elution volume of chymotrypsinogen. It was invariably preceded by a small hump. The immunological activity was spread all over the chromatogram. Areas of immunological activity without any biological activity were located on either side of the biologically active fractions, both in the high molecular weight range (including the void volume) and in the low molecular weight range. The biological LH activity recovered following fractionation on Sephadex G-100 was in close agreement with that loaded, whereas the immunological activity recovered following gel filtration exceeded the loaded activity by a factor of 6–8. In the various physiological states, 11 to 44 % of the total immunological activity recovered was not associated with any biological activity. Furthermore, there was a marked variation in the ratio of biological to immunological activities of those fractions which contained biological activity. It is suggested that the specificity of current RIA methods could be improved significantly by preparing antisera which react only with biologically active LH species.

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