scholarly journals Size characteristics of human leucocyte interferon under reducing and non-reducing conditions

1981 ◽  
Vol 193 (3) ◽  
pp. 947-951 ◽  
Author(s):  
I A Braude ◽  
L S Lin ◽  
W E Stewart
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Apparent Molecular Weight ◽  
Polyacrylamide Gel ◽  
Lower Molecular Weight ◽  
Reducing Conditions ◽  
Size Heterogeneity

Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis was used to characterize human leucocyte interferon (HuIFN-alpha) under reducing and non-reducing conditions. Under non-reducing conditions HuIFN-alpha possesses two size forms, but under reducing conditions (r-HuIFN-alpha) only one is observed. The apparent molecular weight of this one form varies with the concentration of 2-mercaptoethanol used. When r-HuIFN-alpha is permitted to reoxidize the bimodal configuration of HuIFN-alpha is restored. The size heterogeneity of native HuIFN-alpha can be eliminated by mild treatment with NaIO4 [HuIFN-alpha/IO4; Stewart II, Lin, Wiranowska-Stewart & Cantell (1977) Proc. Natl. Acad. Sci. U.S.A. 74, 4200-4204]. The size of the HuIFN-alpha/IO4 increases after treatment with 2-mercaptoethanol (r-HuIFN-alpha/IO4) and the apparent molecular weight of this component also varies with the concentration of 2-mercaptoethanol used. In the case of r-HuIFN-alpha the single peak observed apparently originates from both the higher- and lower-molecular-weight components.

scholarly journals Mapping of the genes controlling high-molecular-weight glutelin subunits of rye on the long arm of chromosome 1R

1984 ◽  
Vol 44 (2) ◽  
pp. 117-123 ◽  
Author(s):  
N. K. Singh ◽  
K. W. Shepherd
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
Translocation Line ◽  
Sds Page ◽  
Chromosome 1R

SUMMARYThe gene(s) controlling the high-molecular-weight glutelin subunits in rye (designated as Glu-Rl) was mapped with respect to the centromere using a 1RL-1DS wheat-rye translocation line and sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). Analysis of 479 seeds from test-crosses between a 1R/1RL-1DS heterozygote and the cultivar India 115, revealed 14·6% aneuploid and 3·95% recombinant progeny. Excluding the aneuploids, this locus was calculated to be 4·65 ± 1·04 cM from the centromere on the long arm of chromosome 1R, which is comparable to the position of the homoeologous loci in wheat and barley.

scholarly journals Purification and characterization of subcomponent C1q of the first component of mouse complement

1981 ◽  
Vol 193 (2) ◽  
pp. 621-629 ◽  
Author(s):  
K Yonemasu ◽  
T Sasaki
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Active Form ◽  
Sodium Dodecyl ◽  
Exchange Chromatography ◽  
Polyacrylamide Gel ◽  
Molecular Weight Range ◽  
Covalently Linked

1. Mouse C1q, a subcomponent of the first component of complement, has been purified in a highly haemolytically active form by a combination of precipitation with EGTA, ion-exchange chromatography and gel filtration. Yields ranged from 3 to 5 mg/200 ml of serum, and the activity of final preparations was in the range of 2 × 10(13)-4 × 10(13) C1q effective molecules/mg. 2. The molecular weight of mouse C1q was 439 500 +/- 1586, as determined by polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. 3. Mouse C1q was shown to be composed of non-covalently linked subunits, all being in the molecular-weight range 45 000-46 000, and three covalently linked chains each having a molecular weight of approx. 23 000 as determined on polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate by using non-covalently and covalently linked subunits of human C1q as markers with known molecular weights calculated theoretically previously [Porter & Reid (1978) Nature (London) 275, 699-704]. 4. Mouse C1q contained hydroxyproline, hydroxylysine, a high percentage of glycine and approx. 9% carbohydrate. The absorption coefficient and nitrogen content of C1q were also determined.

scholarly journals Arterial basement-membrane-like material isolated and characterized from rabbit aortic myomedial cells in culture

1983 ◽  
Vol 211 (2) ◽  
pp. 397-404 ◽  
Author(s):  
L Heickendorff ◽  
T Ledet
Keyword(s):  
Molecular Weight ◽  
Basement Membrane ◽  
High Molecular Weight ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
Type I

Arterial basement-membrane-like material was isolated from rabbit aortic myomedial cell cultures by sonication and differential centrifugation. Isolated basement-membrane-like material was shown to be free of both cellular and matrix contaminants, on the basis of determinations of DNA, RNA, cholesterol, phosphorus and (Na+ + K+)-activated ATPase, combined with electron microscopy. Amino acid analyses showed that arterial basement-membrane-like material was composed of predominantly non-collagenous amino acids. Evaluated by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, reduced basement-membrane-like material comprised six major and about 30 minor components in the Mr range 10 000-600 000. One of the major peptides (Mr 225 000) was disulphide-linked. Periodic acid-Schiff staining of gels indicated that most high-molecular-weight components were glycoproteins. Two-dimensional gel electrophoresis resolved reduced basement-membrane-like material into more than 100 components, with pI from 5 to 7. The disulphide-linked Mr-225 000 peptide appeared heterogeneous, with pI of 5.6-6.0, and was considered to represent fibronectin. All major peptides were of non-collagenous nature, on the basis of their susceptibility to pepsin and resistance to collagenase. Purified myomedial basement-membrane-like material contained collagenous peptides, as indicated by the presence of hydroxyproline and hydroxylysine. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of pepsin-treated and reduced basement-membrane-like material revealed five high-molecular-weight collagenous components appearing in the Mr range 105 000-375 000 relative to type I collagen standards.

scholarly journals Purification and characterization of a plasminogen activator secreted by a pig kidney cell line

1984 ◽  
Vol 219 (3) ◽  
pp. 971-978 ◽  
Author(s):  
M Sudol ◽  
E Reich
Keyword(s):  
Plasminogen Activator ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
Pig Kidney ◽  
Reducing Conditions ◽  
Pig Kidney Cells

The plasminogen activator secreted by calcitonin-treated pig kidney cells was purified, characterized and compared with human urinary urokinase. The purification procedure was based on the following steps: sulphopropyl-Sephadex chromatography, p-aminobenzamidine-Sepharose chromatography, preparative sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and isoelectrofocusing. The purified enzyme was obtained from the conditioned medium with a yield of 13% and a purification factor of 390-fold. Analysis by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis under non-reducing conditions showed one closely spaced doublet with an Mr of 50 000; in the presence of reducing agents, two additional bands of Mr 30 000 and 20 000 appeared. The purified enzyme resembles the 53 000-Mr components of human urinary urokinase in amino acid composition and two-dimensional tryptic peptide maps and in its catalytic properties, and the two enzymes cross-react immunologically with rabbit antibodies raised against either. The enzyme appears to be different from tissue plasminogen activator secreted by HeLa cells.

scholarly journals Anomalous migration of leghaemoglobin on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis

1978 ◽  
Vol 169 (1) ◽  
pp. 251-253 ◽  
Author(s):  
P Lehtovaara
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
Relative Mobility ◽  
Concomitant Decrease ◽  
Helical Content

The estimate of the molecular weight of leghaemoglobin by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis is about 20% too low. This is due to an anomalously high limiting relative mobility. Leghaemoglobin binds 1.4 g of sodium dodecyl sulphate/g of protein with a concomitant decrease in the helical content from 71-72% to 49-51%.

scholarly journals Properties of a 5′-nucleotidase purified from mouse liver plasma membranes

1973 ◽  
Vol 133 (1) ◽  
pp. 189-199 ◽  
Author(s):  
W. H. Evans ◽  
James W. Gurd
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Mouse Liver ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Polyacrylamide Gel ◽  
Tris Buffer ◽  
Nucleotidase Activity

1. Extraction of a mouse liver plasma-membrane fraction with a detergent buffer, N-dodecylsarcosinate–Tris buffer (sarcosyl–Tris buffer), solubilized 90% of the protein and 70% of the 5′-nucleotidase activity. 2. The proteins of the sarcosyl–Tris buffer extract were fractionated by a rate-zonal centrifugation in a sucrose–detergent gradient. The major protein peak sedimented ahead of phospholipids, which mainly remained in the overlay. Glycoproteins were separated ahead of the protein peak. 3. The 5′-nucleotidase activity peak was associated with 5% of the protein applied to the gradient, and contained relatively few protein bands. 4. The 5′-nucleotidase was purified further by gel filtration on Sepharose and Sephadex columns equilibrated with sarcosyl–Tris buffer, to give a single glycoprotein band on sodium dodecyl sulphate–polyacrylamide-gel electrophoresis. The purified enzyme was lipid-free. 5. Electrophoresis in polyacrylamide gels in sarcosyl–Tris buffers showed that the enzymic activity was coincident with the protein band. 6. The molecular weight suggested for the enzyme activity by gel filtration or centrifugation in sucrose gradients was 140000–150000. Sometimes, a minor enzyme peak of lower molecular weight was obtained. 7. Polyacrylamide-gel electrophoresis in sodium dodecyl sulphate indicated that as the polyacrylamide concentration was increased from 5 to 15%, the apparent molecular weight of the enzyme decreased from 130000 to 90000. 8. The evidence that 5′-nucleotidase is composed of two active and similar, if not identical, glycoprotein subunits and the role of detergent in effecting the separation of membrane proteins and glycoproteins are discussed. 9. Substrate requirements, pH optima and the nature of inhibition by an analogue of adenosine diphosphate are reported.

scholarly journals Proteins of the kidney microvillus membrane. Identification of subunits after sodium dodecylsullphate/polyacrylamide-gel electrophoresis

1976 ◽  
Vol 159 (2) ◽  
pp. 395-407 ◽  
Author(s):  
A G Booth ◽  
A J Kenny
Keyword(s):  
Molecular Weight ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Apparent Molecular Weight ◽  
Neutral Endopeptidase ◽  
Polyacrylamide Gel ◽  
British Library ◽  
Major Band ◽  
Microvillus Membrane

The proteins of microvilli prepared from pig kidney were analysed by polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. The typical pattern stained for protein revealed five major bands, four of which also stained for carbohydrate, and about 15 minor bands. For descriptive purposes the bands were designated numerically by their apparent molecular weights (X10(-3). Well-characterized proteins were identified with four of the five major bands. Dipeptidyl peptidase IV, a serine proteinase that may be specifically labelled with di-isopropyl [32P]phosphorofluoridate, was assigned to band 130. Aminopeptidase M was assigned to band 160, though when released from the membrane by a proteinase, this protein comprises three polypeptides each of lower apparent molecular weight than the native enzyme. Neutral endopeptidase can be assigned to band 95 and actin to band 42. The fifth major band (180) is an extrinsic glycoprotein that has not been identified with any microvillus enzyme activity. These four proteins contribute 21% of the microvillus-membrane protein. Kidney microvillus actin was characterized by a variety of properties and was similar to muscle actin. A computer analysis of the gel pattern indicates that it comprises 9.0% of the microvillus protein. Myosin is not present in the microvillus, but another protein associated with band 95, with properties that distinguish it from neutral endopeptidase, was tentatively identified as α-actinin. Alkaline phosphatase was identified as a monomeric polypeptide with an apparent molecular weight of 80000; it is a minor protein of the microvillus and is not discernible as a discrete band in the gel pattern. These and other results permit a model of the organization of the microvillus protein to be suggested. The computer program used has been deposited as Supplementary Publication SUP 50070 (12 pages) at the British Library Lending Division, Boston Spa. Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms given in Biochem. J. (1976) 153,5.

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