Further studies on gel filtration of urinary substances.

Urine specimens from healthy children were concentrated and filtered through Sephadex G-100 and G-25. The eluate was recovered in fractions of 5 ml each. Total hexoses, galactose, total proteins and total uronic acids were determined on each fraction. Total hexoses and galactose were eluted in 6 distinct peaks. The uronic acid containing GAG were eluted in 4 small peaks on G-25 and 1 peak on G-100. This peak is eluted always with high amounts of galactose containing glycoproteins.     

Further studies on hepatic acidic glycosaminoglycans in the Hurler syndrome

Summary Acid GAG were isolated from hepatic tissue from 3 patients with Hurler syndrome and 3 normal controls. Gross elevations in the uronic acid and hexosamine contents were found in Hurler livers compared with the normal ones. The total GAG concentration was significantly increased (about 25fold) in Hurler patients.This work was partially supported by research grants from the Conicet, Argentina.     

Some characteristics of urokinase released in organ culture of human kidney

Summary Plasminogen activator produced in organ culture of human kidney, i.e. in the histotypical arrangement of the tissue, was partially purified by affinity chromatography on para-aminobenzamidine coupled to Sepharose by a 6-carbon spacer, followed by gel chromatography on Sephadex G-100. The molecular weight of 2 active peaks were 27,000 and 52,000 daltons respectively. It was inhibited by DFP and by IgG antiurokinase.This study was supported by grant from the Swedish Medical Research Council (B77-17X-04523-03B).     

Separation of modulator-dependent protein kinase (type I) from cyclic GMP-dependent protein kinase in mouse testes

A new type of enzyme, modulator-dependent protein kinase (type I) (M-PKI), was successfully isolated from the cytosol fraction of mouse testes. It was eluted slightly after the peak of cyclic GMP-dependent protein kinase (G-PK) by Sephadex G-200 gel filtration. Unlike either cyclic AMP-dependent protein Kinase (A-PK) or G-PK, its maximal activity depended exclusively on the presence of crude protein kinase modulators (PKM) or partially purified stimulatory modulator (PKMs).     

Further studies on hepatic acidic glycosaminoglycans in the Hurler syndrome.

Acid GAG were isolated from hepatic tissue from 3 patients with Hurler syndrome and 3 normal controls. Gross elevations in the uronic acid and hexosamine contents were found in Hurler livers compared with the normal ones. The total GAG concentration was significantly increased (about 25fold) in Hurler patients.     

FSH receptor binding inhibiting activity associated with low molecular weight inhibin from sheep,human, rat and chicken

Summary Low molecular weight inhibin (1500 daltons) was obtained from sheep, human, rat and chicken testes by sequential chromatography on Sephadex G-100 and G-25. In addition to its ability to suppress circulating FSH levels in adult castrated male rats, it also exhibits binding inhibition of I¹²⁵hFSH to rat testicular receptors.     

Some characteristics of urokinase released in organ culture of human kidney.

Plasminogen activator produced in organ culture of human kidney, i.e. in the histotypical arrangement of the tissue, was partially purified by affinity chromatography on para-aminobenzamidine coupled to Sepharose by a 6-carbon spacer, followed by gel chromatography on Sephadex G-100. The molecular weight of 2 active peaks were 27,000 and 52,000 daltons respectively. It was inhibited by DFP and by IgG antiurokinase.     

Bombesin-like immunoreactivity in the pancreas of man and other mammalian species

Summary Bombesin-like immunoreactivity has been measured in pancreatic tissues of man (12.4±1.2 pmol/g), pig (15.8±3.2), calf (4.3±0.9), rat (8.5±1.2) and guinea-pig (2.8±0.6) by a specific radioimmunoassay. Gel filtration of the pancreatic extracts revealed 2 major immunoreactive peaks: the earlier peak was eluted in the position of porcine gastrin-releasing peptide, and the later peak was eluted just after the amphibian bombesin standard. Immunocytochemistry demonstrated the presence of bombesin-like immunoreactivity in nerves in the rat pancreas, particularly in the exocrine pancreas, and occasionally in the peri-insular spaces. Isolated rat pancreatic islets were found to contain small quantities of bombesin-like immunoreactivity (0.037±0.003 fmol/islet) suggesting that mammalian bombesin-like peptides may be invovled in the regulation of endocrine as well as exocrine pancreatic secretion.Acknowledgments. We are grateful to Dr. A. V. Edwards, Physiological Laboratory, Cambridge, U. K. for providing the calf tissues, and the British Diabetic Association, U. K. for support.To whom reprint requests should be addressed.     

Partial purification and characterization of acid phosphatase from sporulated oocysts of Eimeria tenella

Acid phosphatase of Eimeria tenella oocysts (Peak II) was purified 77-fold with a recovery of 26% using protamine sulfate precipitation, DEAE-cellulose chromatography and Sephadex G-200 gel filtration. This enzyme occurs in multiple forms as indicated by two peaks which can be separated by DEAE-cellulose chromatography and polyacrylamide gel electrophoresis. The partially purified enzyme has optimal activity at pH 4.5. With p-nitrophenyl phosphate the Km and Vmax values for (Peak II) were 25 mM and 1.57 mumol/min/mg protein, respectively. The enzyme (Peak II) is strongly inhibited by Hg++, Cu++, iodoacetamide, fluoride and molybdate. Tartrate and other divalent metal ions have no effect on enzyme activity. The partially purified Peak II phosphatase is not a glycoprotein as it is not absorbed on concanavalin-A Sepharose and its treatment with bacterial neuraminidase does not alter its elution profile through DEAE cellulose.     

Bradykinin-potentiating peptides from the venom ofAgkistrodon halys blomhqffii

Zusammenfassung Aus dem Gift vonAgkistrodon halys blomhoffii konnten durch Säulenchromatographie an Sephadex G-100, G-25 und CM-Sephadex C-50 fünf Peptidkomponenten isoliert werden, welche die kontrahierende Wirkung von Bradykinin auf den isolierten Meerschweinchendarm signifikant erhöhen.

---

---

Part of this paper was presented at the 41st Annual Meeting of the Japanese Biochemical Society, Tokyo (1968).     

5-Hydroxytryptamine binding to butanol extracts from myelin fragments

Summary The myelin fraction of rat brain stem was treated with butanol-water mixtures, and the extracted proteolipids were separated by Sephadex LH₂₀ column chromatography. 2 peaks of proteolipids eluted in chloroform-methanol 4/1 showed the binding capacity for C¹⁴·5-HT. This finding suggests the necessity of the more careful investigations for the probability of proteolipids as receptor proteins in the central nervous system.     

Isolation and characterization of a high-molecular-weight glycoprotein from the endometrium of porcine uteri

A novel glycoprotein was isolated from the endometrium of porcine uteri. This high-molecular-weight glycoprotein consisted of 25% of protein and 73% of carbohydrate. The carbohydrate composition was quite characteristic in that equimolar N-acetylglucosamine and galactose were major constituents. Its unique nature makes it distinguishable from hitherto-reported glycoproteins.     

Isolation and characterization of a high-molecular-weight glycoprotein from the endometrium of porcine uteri

Summary A novel glycoprotein was isolated from the endometrium of porcine uteri. This high-molecular-weight glycoprotein consisted of 25% of protein and 73% of carbohydrate. The carbohydrate composition was quite characteristic in that equimolar N-acetylglucosamine and galactose were major constituents. Its unique nature makes it distinguishable from hitherto-reported glycoproteins.     

Partial purification and characterization of acid phosphatase from sporulated oocysts ofEimeria tenella

Summary Acid phosphatase ofEimeria tenella oocysts (Peak II) was purified 77-fold with a recovery of 26% using protamine sulfate precipitation, DEAE-cellulose chromatography and Sephadex G-200 gel filtration. This enzyme occurs in multiple forms as indicated by two peaks which can be separated by DEAE-cellulose chromatography and polyacrylamide gel electrophoresis. The partially purified enzyme has optimal activity at pH 4.5. With p-nitrophenyl phosphate the Km and V_max values for (Peak II) were 25 mM and 1.57 mol/min/mg protein, respectively. The enzyme (Peak II) ist strongly inhibited by Hg⁺⁺, Cu⁺⁺, iodoacetamide, fluoride and molybdate. Tartrate and other divalent metal ions have no effect on enzyme activity. The partially purified Peak II phosphatase is not a glycoprotein as it is not absorbed on concanavalin-A Sepharose and its treatment with bacterial neuraminidase does not alter its elution profile through DEAE cellulose.     

Glycosaminoglycans in separated tubules of the guinea-pig and rat kidney medulla

Summary Isolated tubules of the renal medulla of guinea-pig and rat contained glycosaminoglycans. 20–25% of the uronic acids corresponded to hyaluronic acid. In the guinea-pig, chondroitin and dermatan-sulfates accounted for at least 50% of the uronic acids, whereas, in the rat, heparan sulfates comprised 65–70% of them.Supported, in part, by CONICET, Argentina, Mrs N. Ramonda-Becerra assisted in the electrophoretic assays.     

Partial purification of components from fasting human blood serum which stimulate the forward migration of human spermatozoa

Summary Components from fasting human serum that could stimulate the forward migration of human spermatozoa were isolated by gel filtration on Sephadex G-25. The results suggest that these components may be used to enhance sperm forward migration and, hence, pregnancy rate in artificial insemination with husband's semen, (AIH) especially in cases where the sperm forward migration is not optimal.This work was supported by a grant from the World Health Organization.     

Resolution of small G1 and G2 populations of L1210 leukemia by flow microfluorometric analysis aided by velocity sedimentation

Summary L1210 leukemic cells grown in vitro were subjected to kinetic analysis using a flow microfluorometer. A single broad peak was found for the DNA content distribution if unfractionated cells were used; prior fractionation using lg velocity sedimentation allowed the separation of small peaks with smaller (G₁) and larger (G₂) DNA contents from the dominant S phase peak with intermediate DNA content.This work was supported by grant number 5P01CA13053 awarded by the National Cancer Institute, DHEW USA, and by grant num ber RBI 76-1 from the Queen Wilhelmina Fund, The Netherlands.     

Peptide-polysaccharides isolated from the culture medium of the fungus Paracoccidioides brasiliensis pathogenic to man

A peptido-polysaccharide isolated from the culture medium of Paracoccidioides brasiliensis has been studied. The crude product is a mixture of several polysaccharides releasing, by hydrolysis glucose, mannose and galactose. The purified fraction contains mannose (88%) and galactose (12%). This galactomannan has a molecular weight varying between 4.6.10(4) and 2.6.10(4). It is associated with a peptide containing four amino-acids.     

Isolation, from anti-dinitrophenyl sera taken at the beginning of immunization, of a protein combining with the hapten and which is not related to immunoglobulins]

A protein, not antigenically related to the immunoglobulins, has been isolated by affinity chromatography from the serum of rabbits immunized with dinitrophenylated human gamma-globulin. Its concentration in the serum reaches maximum during the first three days which follow the injection. On "Sephadex G-200", it is eluted after the ribonuclease. Its electrophoretic mobility is slower than that of the IgG.     

Glycosaminoglycans in separated tubules of the guinea-pig and rat kidney medulla

Isolated tubules of the renal medulla of guinea-pig and rat contained glycosaminoglycans. 20--25% of the uronic acids corresponded to hyaluronic acid. In the guinea-pig, chondroitin and dermatan-sulfates accounted for at least 50% of the uronic acids, whereas, in the rat, heparan sulfates comprised 65--70% of them.