Bovine Liver Extract Research Articles

The Purification and Properties of β-Galactosidase from Bovine Testes

Abstract A β-galactosidase was purified 600-fold from bovine testes by ammonium sulfate precipitation, acetone fractionation, and affinity chromatography on agarose substituted with terminal thio-β-galactopyranosyl residues. The preparation was devoid of protease, α-fucosidase, α-mannosidase, β-glucosidase, β-glucuronidase, and hyaluronidase activities. A wide variety of compounds containing terminal nonreducing galactose residues were hydrolyzed by the enzyme, including proteoglycans, glycoproteins, gangliosides, disaccharides, and nitrophenylgalactosides. Kinetic studies showed that galactose was released at approximately the same maximal rate from several substrates, but that the Km values differed widely. The enzyme exhibited a high affinity for nitrophenylgalactosides (Km less than 10-4 m). The purified β-galactosidase had a pH optimum of 4.3, required sulfhydryl groups for activity, and was neither stimulated nor inhibited by alkalimetal ions or by EDTA. A single band of β-galactosidase activity was observed after disc gel electrophoresis at several pH values. At each pH value the enzyme activity corresponded in migration to the major protein band. The β-galactosidase eluted as a single peak from Sephadex G-200 (apparent molecular weight 68,000). The enzyme catalyzed the transfer of galactose from p-nitrophenyl-β-galactoside to glucose, N-acetylglucosamine, and N-acetylgalactosamine. Galactose, galactal, galactonolactone, and thio-β-galactopyranosides inhibited the enzyme. Two distinct β-galactosidase activities were shown in bovine liver extracts. One enzyme (or enzymes) (type I) was bound to the affinity column whereas the second enzyme (or enzymes) (type II) passed through the column. While type I β-galactosidase had properties similar to those ascribed to the testicular enzyme, type II β-galactosidase did not hydrolyze lactose or transfer galactose from β-galactosides to glucose.

Bursectomy and Thymectomy of Obese Strain (OS) Chickens with Spontaneous Autoimmune Thyroiditis and Simultaneous Experimental Allergic Encephalomyelitis

Abstract Experimental allergic encephalomyelitis (EAE) was induced in obese strain (OS) chickens with spontaneous autoimmune thyroiditis (SAT). The effects of surgical bursectomy (BE) in ovo on the 18th day of incubation and of thymectomy (TE) on the day of hatching on the development of these two diseases were studied within the same animal. EAE was elicited in 6- and 11-week-old unoperated, thymectomized or bursectomized OS chickens by a single injection of a saline extract of bovine spinal cord and complete Freund's adjuvant, with Pertussis vaccine as a coadjuvant. Unimmunized birds and chickens injected with bovine liver extract served as controls. Neonatal TE led to an increase in the severity of SAT irrespective of the consecutive immunization, but resulted in a decrease of EAE. BE in ovo entailed a significant decrease of SAT, but also a marked suppression of EAE. Thus the bursa dependency of SAT in OS chickens has been confirmed, while both bursa-derived and thymus-derived lymphoid cells appear to be involved in the development of EAE in this strain.

2-Keto-4-hydroxyglutarate Aldolase of Bovine Liver: Purification, criteria of purity, and general properties

Abstract 2-Keto-4-hydroxyglutarate aldolase, which catalyzes the reversible cleavage of 2-keto-4-hydroxyglutarate to yield pyruvate and glyoxylate, has been purified 1300-fold from bovine liver extracts. The purified enzyme appears to be homogeneous when examined by electrophoresis on starch gel or cellulose polyacetate (Sepraphore III) or by chromatography on columns of Sephadex. A molecular weight of 120,000 is obtained with a calibrated column of Sephadex G-200. Two protein bands, both of which are enzymatically active, are observed in polyacrylamide gel electrophoresis. Under certain conditions, two protein peaks are also seen in the ultracentrifuge; the individual components are estimated to have molecular weights of 113,000 and 208,000. The enzyme has a pH optimum of 8.8, is not stimulated by divalent metal ions, and is not inhibited by a number of metal chelating agents. No absolute requirement for added thiol compounds can be shown, but enzymatic activity is strongly inhibited by p-hydroxymercuribenzoate, N-ethylmaleimide, or iodoacetate.

ANTAGONISM OF INSULIN ACTION ON MUSCLE BY THE ALBUMIN-BOUND B CHAIN OF INSULIN.

1. The presence of a substance associated with human albumin that exerts anti-insulin activity on the isolated rat diaphragm has been confirmed. This factor has been removed from albumin, thereby providing a source of non-antagonistic carrier protein. 2. Derivatives of the polypeptide B chain of insulin obtained by chemical scission of the interchain disulphide bonds have been separated by conventional techniques. In the presence of non-antagonistic albumin, the reduced and sulpho-B chain preparations inhibited insulin action on muscle. 3. The B chain resulting from reductive cleavage of insulin by bovine-liver extracts, in association with human albumin, exhibited a comparable anti-insulin effect. 4. It is postulated that the B chain interacts with albumin to enable solubilization of the chain and that inhibition of insulin action on muscle may occur as a result of competition for cellular receptor sites by the B chain. 5. The implication of these findings in relation to a circulating insulin antagonist is discussed.

貝類廢棄部の利用に関する研究-III

A fractionation by treating with activa ?? ed charcoal has been made of growth factors for chicks which were found in the boiled extract of the clam, Venerupis philippinarum. A filtrate fraction which was obtained on treating a slightly acidic solution of the boiled extract with the adsorbent, still possessed growth factors in a con ?? iderable amount. This is not case with bovine liver extract. An eluate from the adsorbate with a dilu ?? e sodium hydroxide solution showed a growth promoting activity nearly equal to that of the filtra ?? e frac ?? ion. The a ?? say of vitamin B12 by Euglena gracilis indica ?? ed that 76 per cent of the vitamin in the extract passed into the non-adsorbed fraction, and that only 0.19 per (e ?? t was found in the eluate. It follows, accordingly, that growth accelerating activity manifested by the eluate must be due to the presence of some unidentifiel growth factor or factors, probably vitamin B15 postulated by TOMIYAMA and YONE2).

肝臓の造血成分に関する研究(第3報)原子爆弾症に対する効果

The present paper deals with the effect of a bovine liver extract upon the recovery in blood-pictures of several serious cases and in blood and bone marrow-picture of one typically serious case. The liver extract used has been prepared by a following procedure; extract an autolyzed liver with water, add ethanol to make its concentration 70 per cent, concentrate the alecoholie solution in vacuo, and dilute a syrup obtained so as to make 1cc of the resultant solution to correspond to five grams fresh liver. The comparison between serious cases administered with the liver extract and those without added it has been made of the changes in hemoglobin concentration, red blood cell count and white blood cell count which were observed during stay in the hospital until 30 Sept., 1945 (55 days elapsed after the bombardment). It will be seen from Table 3 that the liver extract was as much effective as the blood-transfusion in increasing the rate of increase in W. B. C. It is also of interest that a simultaneous treatment with both the liver extract and the blood-transfusion gave rise to an outstanding effect in increasing W. B. C. The examination of blood-and bone marrow-picture of one serious case revealed that the intra-muscular administration of the liver extract was effective in curing a critical stage of a serious type in contrast to some other critical cases which did not receive the liver extract. The change in bone marrow during the treatment with the liver extract is as follows. Nearly complete destruction of bone marrow tissue was observed on 32 nd day after the bombardment; the recovery of R. B. C. system as well as W. B. C. system was under way on 53 rd day (20cc of the liver extract having been administered); mother cells of R. B. C., W. B. C. and blood platelets, and their mature forns were found on 76 th day when 40cc of the liver extract had been administered. From the above results of the clinical examiuations it can be postulated that the liver extract most probably influcence directly on the regeneration or reactivation of bone marrow tissue. Furthermore, the work recently reported by Carter et al. of the effect of vitamin B12 upon leukopenia produced by irradiation of X-ray, seems to suggest that the active principle effective to the regeneration of W. B. C. in the atomic bomb-disease will not be vitamin B12. (From Laboratory of Internal Medicine and of Fishery Chemistry of the Kyushu University)