scholarly journals Tammar Wallaby Plasma Protease Inhibitory (Pi) Proteins

1987 ◽  
Vol 40 (4) ◽  
pp. 355 ◽  
Author(s):  
S D Patterson ◽  
K Bell ◽  
WE Poole
Keyword(s):  
Protease Inhibitor ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Tammar Wallaby ◽  
Island Population ◽  
Sialic Acid Content ◽  
Protein Blotting ◽  
Macropus Eugenii ◽  
Isoelectric Points ◽  
Ph Range

Electrophoretic examination (isoelectric focusing and polyacrylamide gel electrophoresis) of 157 plasmas from a Kangaroo Island population of tammar wallabies (Macropus eugenii) resulted in the identification of five putative condominant protease inhibitor alleles, F, I, M, P and S, which exhibited microheterogeneity due to variable terminal sialic acid content. The frequencies of the five alleles in this popUlation were 0.041(F), 0.682(1), 0.194(M), 0.073(P) and O.OIO(S). The proteins had isoelectric points in the pH range 3.94-4.38, Mr of 60 500 to 66 000 and were identified as protease inhibitors by their abilities to inhibit both trypsin and chymotrypsin. Protein blotting of the denatured proteins demonstrated cross reaction with antiserum to human ai-protease inhibitor.

Plasma protein polymorphisms in the tammar wallaby, Macropus eugenii

1998 ◽  
Vol 46 (2) ◽  
pp. 193 ◽  
Author(s):  
H. Arthur ◽  
K. Bell ◽  
D. W. Cooper
Keyword(s):  
Plasma Protein ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
South Australia ◽  
Backcross Progeny ◽  
Tammar Wallaby ◽  
Vitamin D Binding Protein ◽  
Macropus Eugenii ◽  
Human Proteins ◽  
Protein Polymorphisms

Five populations of the Australian tammar wallaby, Macropus eugenii, from Kangaroo Island, South Australia, and Garden, Abrolhos and Middle Islands and Perup, Western Australia, were examined for plasma protein polymorphisms. Select Kangaroo/Garden Island hybrids and backcross progeny were also included in the study. Vitamin D binding protein (GC), albumin (ALB), transferrin (TF), protease inhibitor (PI), haemopexin (HX), haptoglobin (HP) and immunoglobulin G (IgG) were identified by polyacrylamide gel electrophoresis, pH 7.9, isoelectric focusing, pH 4.2–4.9, and immunoblotting with rabbit antisera to human proteins. Five GC (A, B, C, D, E), two ALB (A, B), two TF (A, B) and five PI (I, J, L, M, P) variants were detected, and limited family studies demonstrated a codominant allelic inheritance for each of the systems.

Biochemical studies of intrauterine components of the tammar wallaby Macropus eugenii during pregnancy

Development ◽  
1981 ◽  
Vol 62 (1) ◽  
pp. 325-338
Author(s):  
Elizabeth J. Thornber ◽  
Marilyn B. Renfree ◽  
Gregory I. Wallace
Keyword(s):  
Protein Concentration ◽  
Specific Binding ◽  
Polyacrylamide Gel Electrophoresis ◽  
Fold Increase ◽  
Tammar Wallaby ◽  
Macropus Eugenii ◽  
Tenfold Increase ◽  
Specific Proteins ◽  
Wet Weight

The in vitro uptake and incorporation of [3H]ui idine by blastocysts of the tammar wallaby showed a 16- and 30-fold increase from day 0 to day 10 after removal of pouch young, respectively. Two of the six non-expanded blastocysts recovered on day 5 showed a tenfold increase in incorporation. During the first ten days after removal of pouch young the diameter of the blastocyst increased threefold. Endometrial exudate from gravid uteri had a higher protein concentration than exudate from nongravid uteri (39·5 ± 0·9 and 32·0 ± 2·0 mg/ml (mean ± s.e.m.), respectively). Endometrial exudates from uteri where the blastocyst was actively growing were found to contain six uterine-specific proteins. These were separated by gradient polyacrylamide gel electrophoresis. Two of the proteins were pre-albumins and the others were larger molecules (M.W. 153000–670000). Two proteins were only present at particular stages of pregnancy: the other four were present at all stages from diapause to birth, in exudate from gravid and nongravid uteri. The specific binding of progesterone and androstenedione to proteins in endometrial exudates or uterine flushings from pregnant wallabies was less than one per cent of the value obtained from day-5 pregnant rabbits. The ability of mouse blastocysts to take up and incorporate [3H]uridine into acidinsoluble material increased threefold in the presence of day-10 endometrial exudates from wallabies. However, this was less than ten percent of the values obtained in the presence of bovine serum albumin. The concentration of calcium in endometrial exudates increased from 23·6 to 45·2 μg/ml during pregnancy; in endometrium it remained at 88·7 μg/g (wet weight) throughout pregnancy, and in plasma it was 53·3 μg/ml. The concentration of zinc in endometrial exudates was 4·5 μg/ml; in endometrium it decreased from 21·8 to 13·3 μg/g (wet weight) during pregnancy and in plasma it was 0·6 μg/ml.

scholarly journals Purification and properties of β-N-acetylhexosaminidase from the mollusc Helicella ericetorum Müller

1978 ◽  
Vol 175 (2) ◽  
pp. 743-750 ◽  
Author(s):  
P Calvo ◽  
A Reglero ◽  
J A Cabezas
Keyword(s):  
Active Site ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Mixed Substrates ◽  
Ph Optimum ◽  
Terrestrial Mollusc ◽  
Buffer Solutions ◽  
Purification And Properties ◽  
Competitive Inhibitors ◽  
Ph Range

1. A beta-N-acetylhexosaminidase was purified 330-fold from the digestive gland of the terrestrial mollusc Helicella ericetorum Müller. 2. Its pH optimum is 4.5 for both beta-N-acetylglucosaminidase and beta-N-acetylgalactosaminidase activities in two buffer solutions; it is fully stable at 37 degrees C for 2h in the pH range 3.8–4.6 and shows one isoelectric point (pH 4.83). 3. The estimated mol.wt. is between 120,000 and 145,000. 4. The enzyme shows an endo-beta-N-acetylhexosaminidase activity on natural substrates such as ovalbumin, ovomucoid, chondroitin 4-sulphate, chitin and hyaluronic acid. 5. Two forms of the enzyme were separated by preparative polyacrylamide-gel electrophoresis. 6. Km and Vmax. for p-nitrophenyl 2-acetamido-2-deoxy-beta-D-glucopyranoside and p-nitrophenyl 2-acetamide-2-deoxy-beta-D-galactopyranoside are 0.43 mM, 30.1 micronmol of p-nitrophenol/min per mg and 0.19 mM, 8.6 micronmol of p-nitrophenol/min per mg respectively. 7. It is inhibited by Hg2+, Fe3+, acetate, some lactones, N-acetylgalactosamine, N-acetylglucosamine and mannose. 8. Mixed-substrates analysis and Ki values for competitive inhibitors indicated that beta-N-acetylglucosaminidase and beta-N-acetylgalactosaminidase activities are catalysed by the enzyme at the same active site.

scholarly journals Body-Surface Compounds in Buckfast and Caucasian Honey Bee Workers (Apis Mellifera)

2014 ◽  
Vol 58 (1) ◽  
pp. 5-15 ◽  
Author(s):  
Aneta Strachecka ◽  
Grzegorz Borsuk ◽  
Jerzy Paleolog ◽  
Krzysztof Olszewski ◽  
Milena Bajda ◽  
...  
Keyword(s):  
Protease Inhibitor ◽  
Body Surface ◽  
Gel Electrophoresis ◽  
Protein Concentration ◽  
Polyacrylamide Gel Electrophoresis ◽  
Chemical Compounds ◽  
The Body ◽  
Surface Chemical ◽  
Surface Compounds ◽  
Honey Bee Workers

Abstract Body-surface chemical compounds were studied in 1-day-old nest workers and foragers both in Buckfast and Caucasian bees. The workers of these two age-castes were sampled twice in each of two consecutive years. Body-surface lipids were determined by means of gas chromatography, with a GCQ mass spectrometer. Protein concentrations and activities on the body surface were examined in bee cuticle rinsings obtained from worker bees according to the methods of Lowry, of Anson, and of Lee and Lin. Protease and protease inhibitor activities were determined. Polyacrylamide gel electrophoresis was performed. Caucasian bees, particularly foragers, had more lipids, but Buckfast bees (two age-castes) had more proteins on their body surfaces. A total of 17 alkane types (C17 - C33), 13 alkene types (C21 - C33), 21 esters (C12 - C32), and a phenol (C14) were detected in both races. Alkene C33 was detected only in Caucasian bees. More alkanes, esters, and phenols were found in Caucasian 1-day-old nest workers and foragers than in these age-castes of Buckfast bees. The protein concentration and protease inhibitor activities were lower in Caucasian bees that had higher protease activities. These values corresponded with specific numbers and widths of the electrophoretic bands.

Studies of the heterogeneity of human pituitary LH by fast protein liquid chromatography

1985 ◽  
Vol 105 (3) ◽  
pp. 405-413 ◽  
Author(s):  
A. Stockell Hartree ◽  
J. B. Lester ◽  
R. C. Shownkeen
Keyword(s):  
Liquid Chromatography ◽  
Sialic Acid ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Binding Activity ◽  
Fast Protein Liquid Chromatography ◽  
Human Pituitary ◽  
Sialic Acid Content ◽  
Receptor Binding Activity

ABSTRACT The Pharmacia fast protein liquid chromatography system was employed to fractionate a purified preparation of human LH (hLH) on the anion exchanger Mono Q at pH 7·8 into 14 sub-fractions. Each of the sub-fractions was characterized by its behaviour on polyacrylamide gel electrophoresis, sodium dodecyl sulphate (SDS) gel electrophoresis, LH receptor binding activity and sialic acid content. All sub-fractions contained sialic acid, were active in binding to LH receptors, and exhibited components typical of hLH subunits on SDS gel electrophoresis. None of the sub-fractions was homogeneous with respect to charge. There is evidence that part of the heterogeneity results from the presence in some molecules of an internal proteolytic cleavage within the β-subunit, and fractions enriched in species containing such cleavages were prepared by this method. J. Endocr. (1985) 105,405–413

scholarly journals Albumin associated with purified pig lymphocyte plasma membrane

1980 ◽  
Vol 192 (1) ◽  
pp. 49-57 ◽  
Author(s):  
M J Owen ◽  
B H Barber ◽  
R A Faulkes ◽  
M J Crumpton
Keyword(s):  
Plasma Membrane ◽  
Serum Albumin ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Membrane Fraction ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Reducing Conditions ◽  
Isoelectric Points ◽  
Pig Serum

Plasma-membrane preparations purified from pig lymphocytes contained a major polypeptide component of mol.wt. about 68 000. This component was identified as pig albumin by the following comparisons with authentic pig serum albumin: (a) co-migration when analysed by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis under reducing and non-reducing conditions; (b) identical isoelectric points; (c) similar “fingerprints” of arginine-containing tryptic peptides; (d) reactivity with anti-(pig albumin) serum. The albumin was bound tightly to the plasma membrane. Biosynthetic labelling of pig lymphocytes under a variety of conditions failed to provide evidence that albumin was synthesized by lymphocytes, suggesting that the plasma-membrane-associated albumin was of extraneous origin. Radiolabelled pig serum albumin, however, failed to bind to the plasma-membrane fraction when added before cell disruption. Although lymphocyte plasma membrane preparations from other species possessed a polypeptide of about 68 000 mol.wt., this was judged not to be albumin on the basis of electrophoretic mobility under non-reducing conditions; also, no polypeptide was precipitated by anti-albumin sera. It is concluded that pig lymphocyte plasma-membrane preparations possess albumin which, although firmly attached, was probably of extraneous origin. This association appeared not to be common to lymphocytes from other species.

scholarly journals Structural and functional microheterogeneity of rat thyroxine-binding globulin during ontogenesis

1992 ◽  
Vol 286 (1) ◽  
pp. 125-130 ◽  
Author(s):  
M Rouaze-Romet ◽  
R Vranckx ◽  
L Savu ◽  
E A Nunez
Keyword(s):  
Binding Properties ◽  
Developmentally Regulated ◽  
Sialic Acid Content ◽  
Thyroxine Binding Globulin ◽  
Binding Characteristics ◽  
Physiological Conditions ◽  
Age Related ◽  
Isoelectric Points ◽  
Major Carrier ◽  
Ph Range

Thyroxine-binding globulin (TBG), the major carrier of thyroid hormones in human and murine sera, is in the rat a developmentally regulated protein, showing a large surge during post-natal growth followed by virtual disappearance in adults. Here we study as a function of age, from the 19-day embryo to 60 days after birth, the structural and binding characteristics of rat TBG microheterogeneity. Serum obtained throughout development, when pre-incubated with 125I-thyroxine (T4), was shown by isoelectric focusing (IEF; pH range 4-5) to contain six labelled isoforms of TBG, with isoelectric points between 4.25 and 4.55. These isoforms differ in their sialic acid content. The relative labelling densities of the isoforms show age-related changes: in neonates, the bulk of T4 is bound to the most alkaline (least sialylated) TBG isoforms; then, with advancing age, it shifts to the most acidic isoforms. To understand whether this progressive transfer of ligand reflects developmental changes in the relative abundance of isoforms, we submitted sera from rats of different ages to crossed immunoelectrofocusing analysis. We demonstrate that the relative proportions of the TBG isoforms remain fairly constant, independent of the level of total TBG. The most acidic forms always represented the majority (approximately 50%), with the most alkaline ones only representing 15% of total TBG. Experiments based on IEF of charcoal-treated sera, supplemented or not with lipidic serum extracts, further demonstrate that the paradoxical low labelling seen in the neonates for the most abundant highly sialylated isoforms is due to inhibition of their binding abilities by liposoluble components, which are particularly concentrated in the sera at the earlier post-natal ages. These studies represent the first analysis of concentration versus binding functions of rat TBG isoforms in the physiological conditions of normal ontogeny. Our results point to an important influence for the serum environment on the binding properties of TBG isoforms. The physiological significance of such interactions remains to be clarified.

scholarly journals Molecular forms of myeloperoxidase in human plasma

1986 ◽  
Vol 237 (2) ◽  
pp. 559-565 ◽  
Author(s):  
R L Olsen ◽  
T K Steigen ◽  
T Holm ◽  
C Little
Keyword(s):  
Ion Exchange ◽  
Human Plasma ◽  
Gel Electrophoresis ◽  
Large Subunit ◽  
Polyacrylamide Gel Electrophoresis ◽  
Immunoaffinity Chromatography ◽  
Molecular Forms ◽  
Active Components ◽  
Protein Blotting ◽  
Catalytically Active

A radioimmunoassay for myeloperoxidase was established with the use of affinity-purified anti-(human myeloperoxidase) immunoglobulins. By the use of ion-exchange followed by immunoaffinity chromatography a preparation of immunoreactive, catalytically active myeloperoxidase was obtained from fresh human plasma. In non-denaturing gel electrophoresis, the plasma preparation showed about four catalytically active components of mobility very similar to that of the granulocyte enzyme. SDS/polyacrylamide-gel electrophoresis combined with protein blotting showed that the two polypeptides of strongest antigenicity in the plasma preparation corresponded in Mr to the large and the small subunits of the granulocyte enzyme. In addition, the plasma preparation contained a higher-Mr immunoreactive polypeptide, possibly a precursor form of the enzyme, together with another of Mr similar to that of the large subunit of eosinophil peroxidase.

Characterization of the protein Z–dependent protease inhibitor

Blood ◽  
2000 ◽  
Vol 96 (9) ◽  
pp. 3049-3055 ◽  
Author(s):  
Xin Han ◽  
Ryan Fiehler ◽  
George J. Broze
Keyword(s):  
Protease Inhibitor ◽  
Gel Electrophoresis ◽  
Thrombin Generation ◽  
Proteinase Inhibitors ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Factor Xa ◽  
Polyacrylamide Gel ◽  
Protein Z ◽  
Factor Xia

Abstract Protein Z-dependent protease inhibitor (ZPI) is a 72-kd member of the serpin superfamily of proteinase inhibitors that produces rapid inhibition of factor Xa in the presence of protein Z (PZ), procoagulant phospholipids, and Ca++ (t1/2 less than 10 seconds). The rate of factor Xa inhibition by ZPI is reduced more than 1000-fold in the absence of PZ. The factor Xa–ZPI complex is not stable to sodium dodecyl sulfate–polyacrylamide gel electrophoresis, but is detectable by alkaline–polyacrylamide gel electrophoresis. The combination of PZ and ZPI dramatically delays the initiation and reduces the ultimate rate of thrombin generation in mixtures containing prothrombin, factor V, phospholipids, and Ca++. In similar mixtures containing factor Va, however, PZ and ZPI do not inhibit thrombin generation. Thus, the major effect of PZ and ZPI is to dampen the coagulation response prior to the formation of the prothrombinase complex. Besides factor Xa, ZPI also inhibits factor XIa in the absence of PZ, phospholipids, and Ca++. Heparin (0.2 U/mL) enhances the rate (t1/2 = 25 seconds vs 50 seconds) and the extent (99% vs 93% at 30 minutes) of factor XIa inhibition by ZPI. During its inhibitory interaction with factor Xa and factor XIa, ZPI is proteolytically cleaved with the release of a 4.2-kd peptide. The N-terminal amino acid sequence of this peptide (SMPPVIKVDRPF) establishes Y387 as the P1 residue at the reactive center of ZPI. ZPI activity is consumed during the in vitro coagulation of plasma through a proteolytic process that involves the actions of factor Xa with PZ and factor XIa.

Studies of the Metabolism of Asialotransferrins: Evidence that Transferrin Does not Undergo Desialylation in Vivo

1974 ◽  
Vol 46 (6) ◽  
pp. 763-774
Author(s):  
K.-L. Wong ◽  
P. A. Charlwood ◽  
M. W. C. Hatton ◽  
E. Regoeczi
Keyword(s):  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Biological Significance ◽  
Polyacrylamide Gel ◽  
Fractional Catabolic Rate ◽  
Sialic Acid Content ◽  
Catabolic Rate ◽  
Bovine Transferrin

1. Experiments are reported which aimed at determining whether transferrin loses sialyl residues from the carbohydrate side-chains during the biological lifetime of the molecule. To explore this possibility, transferrin fractions of relatively high sialic acid content (referred to as sialotransferrin) were prepared from purified rabbit and bovine transferrin by preparative polyacrylamide-gel electrophoresis. After labelling with 125I, the preparations were injected into a group of three rabbits each. From the plasma samples obtained between 1 h and 6–8 days after injection, transferrin was partially purified, mixed with 131I-labelled asialotransferrin of the corresponding species and run in preparative polyacrylamide-gel electrophoresis. In each specimen examined, the 125I radioactivity migrated ahead of the marker asialotransferrin, and no portion of the dose was detected with the electrophoretic mobility of asialotransferrin. 2. Evidence is presented that bovine transferrin desialylated in vitro remains detectable in the plasma of rabbits for intervals which are comparable with those found in previous studies with rabbit asialotransferrin. 3. A mathematical model is described for the computation of asialo- to sialotransferrin radioactivity ratios in the plasma, continuous desialylation of pulse-injected sialotransferrin being assumed. Calculations were made at various hypothetical rates of desialylation. 4. On the basis of the experimental data and the model it is concluded that transferrin (both rabbit and bovine) is not subjected to systematic desialylation in rabbits. Random desialylation of some transferrin could take place at rates less than 5% of the fractional catabolic rate of transferrin, which would be without any biological significance.

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