摘要:

Human liver NAD+-dependent aldehyde dehydrogenase oxidizes several aldehydes which differ in the extent of their hydration in aqueous solution. Results of experiments indicate that the free carbonyl form is the operative substrate. When nonenzymatic hydration of acetaldehyde was slower than the enzymatic oxidation reaction, acetaldehyde which initially was anhydrous was completely oxidized at an almost linear rate. By contrast, oxidation of acetaldehyde already in equilibrium with hydrate was biphasic. An initial rapid reaction accounted for approximately half the total aldehyde,i.e. the free aldehyde. During the second stage the acetaldehyde, initially in hydrate form, was oxidized at a reduced rate expected for a prior rate-limiting dehydration step. Small amounts of hydroxylamine, capable of reacting rapidly with free formaldehyde, caused transient inhibition of oxidation of aqueous formaldehyde (which is almost completely hydrated). Furthermore, Michaelis constants for crotonaldehyde and benzaldehyde (which are largely unhydrated in aqueous solution) were very low.

ISSN:0008-4018    

DOI:10.1139/o71-001

出版商:NRC Research Press    

年代:1971

数据来源: NRC

摘要:

Fluorescence was measured in the hydrolysates of several proteins and peptides of known amino acid composition. The most intense fluorescence (emission maximum at 445 nm) was found in acid, but not enzymic, hydrolysates of tryptophan-rich proteins; nine fluorescent fractions were resolved from acid hydrolysates of these proteins by gel filtration. Fluorescence of substances in tryptophan-containing proteins is largely eliminated by reduction, but little affected by oxidation or ultraviolet irradiation. The substances are thermostable and apparently are not amino acids.Tyrosine, tyrosine-containing peptides, and tryptophan-free proteins yielded fewer fluorescent products than did tryptophan-rich proteins. The number and amount of fluorescent derivatives produced from tyrosine during acid hydrolysis depends on the particular protein; also, intra- or intermolecular interactions involving tyrosine or its degradation products appear to influence the nature of fluorescence in protein hydrolysates.The present findings suggest possible quantitation of the tryptophan content of proteins by fluorometric measurement of tryptophan derivatives produced by acid hydrolysis.

ISSN:0008-4018    

DOI:10.1139/o71-002

出版商:NRC Research Press    

年代:1971

数据来源: NRC

摘要:

The activity of hyaluronidase per milligram protein was found to be significantly greater in subcellular fractions from kidneys of rats injected 12 or 18 h previously with 10 mg hydrocortisone than it was in control rats. The difference was greater at 12 h than at 6 or 18 h, and at 12 h those fractions containing lysosomes showed the greatest increase. A higher specific activity of hyaluronidase in some subcellular fractions from liver and spleen was also found after hydrocortisone. Cycloheximide, given 5 h after hydrocortisone, abolished the increase in hyaluronidase measured at 12 h. However, actinomycin D givenbefore hydrocortisone caused only a slight decrease in the effect on hyaluronidase in kidney homogenates.

ISSN:0008-4018    

DOI:10.1139/o71-003

出版商:NRC Research Press    

年代:1971

数据来源: NRC

摘要:

A simple procedure for partial purification of a DNA polymerase from immature testes of salmon (Oncorhynchus nerka) in which 20% glycerol and 2 mMdithiothreitol are used to protect the enzyme is described. The enzyme has a specific activity of 17–45 nmoles of deoxyribonucleoside triphosphate (dTTP) incorporated into DNA per milligram protein per hour at 25 °C. It has a specific requirement for Mg2+or Mn2+, for four deoxyribonucleoside triphosphates, and for a DNA primer. The latter is best supplied by a DNA duplex that has undergone limited digestion with pancreatic DNase or by polydeoxyadenylate polydeoxythymidylate alternating copolymer (poly d(A–T)) that has been similarly treated. Native and heat-denatured DNA are poor primers. The enzyme is strongly inhibited by pancreatic DNase and pyrophosphate, less strongly by actinomycin D, and is markedly stimulated by EDTA. Addition of KCl and NaCl causes inhibition that at 100 mMconcentration is 70% with KCl and 80% with NaCl. The enzyme loses 50% of its activity in 15 min at 35 °C and in 3 h at 30 °C. At 30 °C glycerol (47%), DNA, and bovine serum albumin (2 mg/ml) retard loss in activity. With poly d(A–T) primer, incorporation of α-32P-dATP into DNA proceeds for only a short time and then ceases unless dTTP is present, when incorporation continues. When the products formed in such reactions are subjected to nearest-neighbor frequency analysis only 3′-dTMP possesses radioactivity. Polydeoxyguanylate polydeoxycytidylate linear polymer has no priming activity. The product of the DNA polymerase formed a band in a buoyant density gradient of CsCl at a density similar to that of the DNA primer.

ISSN:0008-4018    

DOI:10.1139/o71-004

出版商:NRC Research Press    

年代:1971

数据来源: NRC

摘要:

A 1:1 molar stoichiometry has been found for the noncovalent bonding of proflavine with δ-chymotrypsin. The dissociation constant at pH 7.6 is 0.025 mMwhich is similar to that found by Glazer for the reaction of proflavine with α-chymotrypsin. The difference spectrum produced by the proflavine–δ-chymotrypsin interaction has been studied as a function of pH. It would appear that a group with pKof approximately 6.5 is involved.

ISSN:0008-4018    

DOI:10.1139/o71-005

出版商:NRC Research Press    

年代:1971

数据来源: NRC

摘要:

14C-15α-hydroxyandrostenedione was perfused through two human placentas at the time of Caesarean section. The aromatization of 15α-hydroxyandrostenedione has been confirmed as a pathway for the placental production of 15α-hydroxyestrone and 15α-hydroxyestradiol in the human term placenta.

ISSN:0008-4018    

DOI:10.1139/o71-006

出版商:NRC Research Press    

年代:1971

数据来源: NRC

摘要:

We have partially characterized and localized two previously reported deoxyribonucleases from the rat small intestine. After separation of the crypt cells and muscle (the deep layer) from the villus cells (the superficial layer), the latter was found to contain a deoxyribonuclease I with a pH optimum around 6, and a molecular weight of 32 000 – 35 000. It was activated by Co2+, Mg2+, and Mn2+. Ligation of the pancreatic duct reduced the activity in intestinal extracts to about one-third of control levels. A deoxyribonuclease II with a pH optimum of 5.3–5.4 was found associated with the continuously dividing intestinal crypt cells. It was inhibited by Mg2+and activated by EDTA. Ligation of the pancreatic duct was without effect on this enzyme. The deoxyribonuclease I is probably largely extracellular and serves a digestive function while the deoxyribonuclease II probably is related to intracellular DNA metabolism.

ISSN:0008-4018    

DOI:10.1139/o71-007

出版商:NRC Research Press    

年代:1971

数据来源: NRC

摘要:

Egg yolk lipoproteins were separated into four major density classes by conventional ultracentrifugation, and each class was resolved further by thin-layer chromatography on hydroxylapatite. The plates were developed in special saturation chambers with phosphate buffers of 0.4, 0.6, 1.2, and 2.0 Mfor 2 h, and bands were located by exposure to iodine vapor or by spraying with ninhydrin. The low density fraction and the low density fraction of the granule each gave two subfractions, while the phosvitin–lipovitellin fraction yielded three components and the water-soluble fraction four components. The additional resolution apparently was due to differences in lipid and protein composition and structure, as well as to the content of protein-bound phosphorus. The described separations offer special advantages for the study of the lipid parts of the lipoprotein complexes.

ISSN:0008-4018    

DOI:10.1139/o71-008

出版商:NRC Research Press    

年代:1971

数据来源: NRC

摘要:

Four major density classes of lipoproteins were isolated from hen's egg yolk by conventional ultracentrifugation and each class was subjected to thin-layer chromatography on hydroxylapatite. Lipid extracts were obtained from the total egg yolk, the density classes recovered at each stage of centrifugation and from the fractions resolved on hydroxylapatite. The proportions of the lipid classes in the total lipid extracts were determined by direct gas chromatography. The molecular species of the glycerides and phospholipids were identified by means of specific enzymic hydrolyses following isolation of the lipid classes by thin-layer chromatography.The results indicate that the major differences in the lipid composition of the egg yolk lipoproteins are confined to variations in the proportions of cholesterol, triglycerides, and phospholipids. There are lesser differences in the proportions of the various phospholipid classes among the different lipoproteins. All lipoprotein classes, except the water-soluble fraction, contained the same species of each phospholipid and triglyceride. The lipids of the water-soluble fraction were more saturated than those of the other lipoproteins.

ISSN:0008-4018    

DOI:10.1139/o71-009

出版商:NRC Research Press    

年代:1971

数据来源: NRC

摘要:

Enzymatic assays for CMP-sialic acid: glycoprotein sialyltransferase and UDP-N-acetylglucosamine: glycoproteinN-acetylglucosaminyltransferase were performed on crude homogenates of three Morris hepatomas (7777, 7800, and 5123D), and on liver homogenates from the host animals and normal Buffalo strain rats. It was found that sialyltransferase activities were greatly decreased in the most rapidly growing tumor (hepatoma 7777) and were decreased to a lesser extent in the more slowly growing hepatoma 7800; enzyme activities in hepatoma 5123D, another relatively slow growing tumor, were not significantly different from control values. Sialyltransferase activities were significantly elevated in the livers of all the tumor-bearing animals and were especially high in the livers of animals carrying hepatoma 7777; these elevations may be related to increased plasma glycoprotein synthesis by liver secondary to the inflammatory stimulus generated by the tumors. In contrast to the sialyltransferase analyses,N-acetylglucosaminyltransferase activities in tumor homogenates were very similar to control values for all three hepatomas. When the data are expressed as ratios of sialyltransferase activity toN-acetylglucosaminyltransferase activity, two of the three tumors show highly significant decreases of this ratio compared to either control or host livers. Since these glycosyltransferases have previously been shown to be located in the Golgi apparatus of normal rat liver where they function in the biosynthesis of glycoproteins, the above results have been interpreted to indicate a shift in the function of the Golgi apparatus in certain Morris hepatomas as compared to normal livers. Finally, glycosyltransferase assays and electron microscopy have been used to demonstrate the feasibility of preparing Golgi-enriched fractions from all three hepatomas by methods previously applied to normal rat liver.

ISSN:0008-4018    

DOI:10.1139/o71-010

出版商:NRC Research Press    

年代:1971

数据来源: NRC