DNA Polymerase Activity Associated with Endogenous Template: Release by Ribonuclease Treatment

1975 ◽  
Vol 53 (5) ◽  
pp. 574-582
Author(s):  
Francesco Moranelli ◽  
P. E. Penner
Keyword(s):  
Molecular Weight ◽  
Dna Polymerase ◽  
Gel Filtration ◽  
Polymerase Activity ◽  
Void Volume ◽  
Lower Molecular Weight ◽  
Single Peak ◽  
Gel Filtration Chromatography ◽  
Thymus Extract ◽  
Concomitant Decrease

A ribonuclease-sensitive DNA polymerase, which uses an endogenous template, is detectable in the 39 000 g supernatant of a rat thymus homogenate, and appears as a single peak of activity in the void volume after Sephadex G 150 or G 200 gel filtration chromatography. Native and "activated" DNA-dependent DNA polymerase activities coincide with the endogenous-templated polymerase activity. Treatment of the thymus extract with ribonuclease(s) prior to gel filtration chromatography yields two other peaks of activity in addition to the void volume peak. The appearance of the two lower molecular weight peaks of activity is accompanied by a concomitant decrease in the endogenous-templated activity. The effect of ribonuclease is specific and cannot be reproduced by a similar deoxyribonuclease treatment.

scholarly journals Metalloproteinases from rabbit bone culture medium degrade types IV and V collagens, laminin and fibronectin

1981 ◽  
Vol 199 (3) ◽  
pp. 807-811 ◽  
Author(s):  
G Murphy ◽  
T E Cawston ◽  
W A Galloway ◽  
M J Barnes ◽  
R A D Bunning ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Culture Medium ◽  
Type Iv Collagen ◽  
Gel Filtration ◽  
Lower Molecular Weight ◽  
Bone Culture ◽  
Gel Filtration Chromatography ◽  
Type Iv ◽  
Rabbit Bone ◽  
Type V

Gel-filtration chromatography of culture medium from rabbit bone explants separates three latent metalloproteinases with activities against collagen, proteoglycan and gelatin respectively. The fractions degrading proteoglycan also degrade laminin, fibronectin and the polymeric products of pepsin-solubilized type IV collagen and can also solubilize insoluble type IV collagen. The fractions degrading gelatin are capable of degrading solubilized type V and 1 alpha,2 alpha,3 alpha (cartilage) collagens, as well as the lower-molecular-weight products of pepsin-solubilized type IV collagen. All activities can be inhibited by 1,10-phenanthroline and occur in either partially or totally latent forms that can be activated by 4-aminophenylmercuric acetate.

Soluble Fibrin Complexes and Fibrinogen Heterogeneity in Diabetes mellitus

1980 ◽  
Vol 44 (03) ◽  
pp. 130-134 ◽  
Author(s):  
E B Tsianos ◽  
N E Stathakis
Keyword(s):  
Diabetes Mellitus ◽  
Molecular Weight ◽  
Plasma Proteins ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Lower Molecular Weight ◽  
Polyacrylamide Gels ◽  
Soluble Fibrin ◽  
Α2 Macroglobulin ◽  
Patients With Diabetes

SummaryThe presence of soluble fibrin complexes (SFC) measured by gel filtration of plasma on 4% agarose columns, fibrinogen heterogeneity on 3.5% SDS-polyacrylamide gels and the concentrations of several plasma proteins were evaluated in 39 patients with diabetes mellitus (DM) and 19 matched control subjects. A small but significant increase of SFC was found in DM (p<0.01). On individual basis 51.2% of the patients had increased SFC (>M + 2 SD of the controls). Polyacrylamide gel electrophoresis of the SFC showed no evidence of cross-linking or proteolysis. Plasma clots formed in the presence of EDTA and trasylol were analysed in SDS-polyacrylamide gels in a normal and two lower molecular weight fibrin bands (band I, II, III). The percentage of band I fibrinogen was in diabetics (65.3 ± 4.7%) lower than that of the controls (71.8 ± 4.5%) (p < 0.01). Fibrinogen levels, antithrombin III, α1-antitrypsin, α2-macroglobulin and plasminogen were significantly increased in DM. We suggest that in DM there is an enhancement of intravascular fibrin formation and accelerated fibrinogen degradation to lower molecular weight forms.

scholarly journals High Molecular Weight Factor VIII Coagulant Activity in Cryoprecipitate and Polyethylene Glycol Precipitates

1977 ◽  
Author(s):  
K. A. Rickard ◽  
T. Exner ◽  
H. Kronenberg
Keyword(s):  
Molecular Weight ◽  
Polyethylene Glycol ◽  
Factor Viii ◽  
High Molecular Weight ◽  
Gel Filtration ◽  
Lower Molecular Weight ◽  
Human Antibody ◽  
High Molecular Weight Material ◽  
Coagulant Activity ◽  
Molecular Weight Form

Gel filtration of human plasma cryoprecipitate on Sepharose 2B indicated the molecular weight of factor VIII coagulant activity (VIIIc) to be significantly greater than that found in antihaemophilic concentrate. Polyethylene glycol at 3% concentration precipitated approximately half of the VIIIc from cryoprecipitate. This activity eluted as high molecular weight material on gel filtration. The addition of more polyethylene glycol to a concentration of 8% precipitated most of the remaining VIIIc from cryoprecipitate. This activity appeared to be of significantly lower molecular weight, approximately corresponding in elution volume to that observed for antihaemophilic concentrate. The possibility that an antibody to VIIIc generated in a patient treated with cryoprecipitate might be directed against the higher molecular weight form of factor VIII was investigated. However, no significant differences between the higher and lower molecular weight forms of factor VIII either in stability or in reactivity with human antibody to factor VIII were found.

scholarly journals Difference in Thrombolytic Effect between Higher and Lower Molecular Weight Forms of Urokinase

1977 ◽  
Author(s):  
T. Suyama ◽  
M. Nishida ◽  
Y. Iga ◽  
R. Naito
Keyword(s):  
Molecular Weight ◽  
Gel Filtration ◽  
Lower Molecular Weight ◽  
Dissolution Time ◽  
Thrombolytic Activity ◽  
Lysis Time ◽  
Thrombolytic Effect ◽  
Approximate Molecular Weight ◽  
Made In

Urokinase (UK) from human urine has been widely used for thrombolytic therapy in Japan. However, commercially available preparations are not identical but consist of mainly two forms of UK with higher and lower molecular weight (H-UK and L-UK) . An attempt was made in this report to compare thrombolytic activity of H-UK with that of L-UK on artificial thrombi produced from human blood by a modification of Chandler’s loop method, which was somehow comparable to the situation in vivo. Two active forms of UK were purified from crude preparation by gel filtration. The approximate molecular weight of the H-UK was 54,000 and of the L-UK 34,000. The potency of UK was determined by “two-stage lysis time method” and expressed by International unit(lU). Thrombolytic activity measured by Chandler’s method was calculated as % lysis of the control thrombus that was formed in the abscence of UK.As a result, thrombus-dissolution time of H-UK was much shorter than that of L-UK. Furthermore, concentration of H-UK (IU/ml blood) necessary to induce 50% lysis was approximately one half lower than that of L-UK. The similiar results were obtained on artificial thrombi from the blood of dog, rat and rabbit. The data suggest that H-UK seems to be more effective on treatment of thromboembolicdisorders as compared to L-UK in terms of the same IU basis.

scholarly journals Complex Formation of Covalently Crosslinked Fibrin Oligomers with Agarose Coupled Fibrinogen and Fibrin

1977 ◽  
Author(s):  
R. von Hugo ◽  
R. Hafter ◽  
A. Stemberger ◽  
H. Graeff
Keyword(s):  
Molecular Weight ◽  
Complex Formation ◽  
Affinity Chromatography ◽  
High Molecular Weight ◽  
Calcium Chloride ◽  
Gel Filtration ◽  
Void Volume ◽  
Soluble Fibrin ◽  
Derivatives Of

Crosslinked high molecular weight derivatives of fibrin (fibrinoligomers) were observed during intravascular coagulation. It was the purpose of this study to investigate the complex formation of fibrin oligomers with fibrinogen and fibrinmonomer. Fibrinogen coupled to agarose (Fg-ag) as well as fi-brinmonomer coupled to agarose (Fm-ag) was used as substrate. Soluble oligomers of fibrin were produced by incubating fibrinogen with thrombin, calcium-chloride, cystein and F XIII. They were separated from fibrinogen by gel filtration. Γ-dimers were demonstrated in fractions from the void volume and the shoulder prior to the fibrinogen peak. These fractions were subjected to affinity chromatography. Crosslinked oligomers of fibrin were not adsorbed on Fg-ag, yet adsorption occured on Fm-ag. This indicates that fibrin oligomers have no affinity to fibrinogen, yet readily form complexes with fibrin. This could mean that in vivo they compete with fibrinogen for the fibrinmonomer part of soluble fibrin monomer complexes, and hence have a tendency to increase in size.

scholarly journals Adsorption of F VIII on Aluminium Hydroxide

1977 ◽  
Author(s):  
M. J. Seghatchian ◽  
T. Barrowcliffe ◽  
M. Miller-Andersson
Keyword(s):  
Molecular Weight ◽  
High Purity ◽  
Aluminium Hydroxide ◽  
Gel Filtration ◽  
Procoagulant Activity ◽  
Void Volume ◽  
Low Molecular Weight ◽  
Two Stage ◽  
Volume Fractions ◽  
High Purification

Adsorption of plasma by A1(OH)3 is a requirement for the two stage assay of F VIII. It is generally accepted that factors II, VII, IX and X are removed by the procedure, while factors V and VIII are unaffected. Following gel filtration of a F VIII concentrat on Sepharose 4 B F VIII:c was found in the low molecular weight area, as well as in the void volume as expected. This activity was found with both one and two stage techniques. After adsorption of the fractions with Al(OH)3 to eliminate the non F VIII procoagulant activity F VIII:c disappeared from the void volume fractions and was much reduced in the low molecular region. F VIII: R Ag was also removed from these fractions by A1(OH)3 adsorption. After adsorption of fractions in the presence of hemophilia plasma clotting activity remained in both regions suggesting the presence of true F VIII activity. Thus at concentration of 1 IU of F VIII:c per ml, a low purity preparation was unaffected by A1(OH)3 adsorption whereas both antigen and clotting activity of a high purity concentrate were conciderably reduced. Addition of 5 % albumin to the high purity preparation prevented this adsorption. It is concluded that under conditions of high purification F VIII:c can be adsorbed preferentially on A1(OH)3 and this appears to be due to removal of F VIII:R Ag.

Gel Filtration of Plasma from Rabbits Injected with Honomeric DES-AB 125I-Fibrin and 131I–Fibrinogen

1979 ◽  
Author(s):  
I. Kröhnke ◽  
I. Hahn ◽  
W. Krell ◽  
G. Müller-Berghaus
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Gel Filtration ◽  
Void Volume ◽  
Molecular Configuration ◽  
Chromatographic Behaviour ◽  
Fibrin Monomer ◽  
Deutsche Forschungsgemeinschaft

We intended to study the chromatographic behaviour of soluble des-AB fibrin prepared in vitro and injected into rabbits. To prepare des-AB 1251-fibrin, purified rabbit 125I-fibrinogen was clotted by thrombin and the formed clot dissolved in Tris-buffered 3 M urea. Gel filtration of des-AB fibrin in urea through sepharose-CL 6B columns equilibrated with buffered 3 M. urea revealed monomeric fibrin. Rabbits received 131I-fibrinogen and 5 min later monomeric des-AB 125I-fibrin in urea. 30 min after injection blood was drawn and the plasma obtained filtered through sepharose-CL 6B columns eguilibrated with buffered plasma. At 20°C as well as at 37°C des-AB 125I-fibrin was eluted in the void volume in front of the 131I-fibrinogen peak. The data demonstrate that monomeric des-AB 125I-fibrin in urea injected into rabbits remains soluble in plasma. Possibly, monomerJ fibrin is converted to circulating soluble high-molecular weight fibrin aggregates or fibrin monomer changes its molecular configuration, thus showing a different gel filtration behaviour.(Supported by the Deutsche Forschungsgemeinschaft).

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.

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