scholarly journals The molecular weights of two forms of carbamoyl phosphate synthase from rat liver

1972 ◽  
Vol 127 (3) ◽  
pp. 503-508 ◽  
Author(s):  
R. Virden
Keyword(s):  
Molecular Weight ◽  
Concentration Dependence ◽  
Rat Liver ◽  
Chemical Equilibrium ◽  
High Speed ◽  
Carbamoyl Phosphate ◽  
Molecular Weights ◽  
A Value ◽  
Equilibrium Sedimentation ◽  
Phosphate Synthase

1. N-Acetylglutamate-dependent carbamoyl phosphate synthase from rat liver was centrifuged in sucrose density gradients. The concentration-dependence of s was consistent with a chemical equilibrium existing between the 11S and 7.5S forms of the enzyme. 2. Under conditions favouring the 11S form, the properties of the enzyme in ultra-short-column equilibrium experiments suggest a molecular weight of 316000±42000 for the 11S form. 3. Under conditions favouring the 7.5S form, high-speed equilibrium-sedimentation measurements gave a value of 160000±10000 as the molecular weight of the 7.5S form of the enzyme.

scholarly journals Inhibition of carbamoyl-phosphate synthase (ammonia) by Tris and Hepes. Effect on Ka for N-acetylglutamate

1987 ◽  
Vol 243 (1) ◽  
pp. 273-276 ◽  
Author(s):  
P Lund ◽  
D Wiggins
Keyword(s):  
Rat Liver ◽  
Phosphate Buffer ◽  
Active Enzyme ◽  
Urea Synthesis ◽  
Carbamoyl Phosphate ◽  
A Value ◽  
Phosphate Synthase

The apparent Ka for N-acetylglutamate of rat liver carbamoyl-phosphate synthase is 11 microM in phosphate buffer, a value 10-fold lower than reported in other buffer systems. Tris and Hepes inhibit competitively with N-acetylglutamate. The proportion of carbamoyl-phosphate synthase in the active enzyme-acetylglutamate complex in vivo may be higher than previous calculations suggest, which re-opens the question of the involvement of N-acetylglutamate in the regulation of urea synthesis.

Physicochemical and chemical studies on wheat embryo ribosomal proteins

1968 ◽  
Vol 46 (4) ◽  
pp. 373-380 ◽  
Author(s):  
Fred H. Wolfe ◽  
Cyril M. Kay
Keyword(s):  
Molecular Weight ◽  
Ribosomal Proteins ◽  
High Speed ◽  
Polyacrylamide Gel Electrophoresis ◽  
Average Molecular Weight ◽  
Optical Rotatory Dispersion ◽  
Random Coil ◽  
Wheat Embryo ◽  
Molecular Weights ◽  
A Value

The physical heterogeneity of unfractionated wheat embryo ribsomal proteins, prepared by the glacial acetic acid method of Waller and Harris, has been investigated in 8 M urea −10−3 M dithio-threitol solutions of low pH (4.5). Sedimentation–diffusion measurements resulted in a weight average molecular weight of 29 000 ± 2 500, with no obvious evidence of heterogeneity. High-speed membrane osmometry was employed to establish the number average molecular weight of this system as 24 500 ± 1 000. The disparity in molecular weight averages suggests some size heterogeneity, and statistical analysis based on the two average molecular weights resulted in a calculated range of molecular weights for wheat embryo ribosomal proteins from 15 000 to 34 000 a.m.u. Charge differences, reflecting presumably primary structure differences, also exist among the members of this class, since about 26 different bands were resolved on polyacrylamide gel electrophoresis. The weight intrinsic viscosity of the ribosomal proteins in 8 M urea solutions was established as 0.273 dl/g, a value considerably larger than most globular proteins, suggesting that a major portion of their polypeptide chains are unfolded in this solvent. This conclusion was substantiated by optical rotatory dispersion measurements on this system, which resulted in a dispersion constant, λc, of 213 m μ, a value typical of that of the random coil. Amino acid and N-terminal analyses are also reported for this system, and comparisons of both chemical and physicochemical parameters are made with ribosomal proteins of other sources.

Measurement of the Isothermal Volume Dilation Accompanying the Unilateral Extension of Rubber

1959 ◽  
Vol 32 (2) ◽  
pp. 428-433
Author(s):  
Fred G. Hewitt ◽  
Robert L. Anthony
Keyword(s):  
Molecular Weight ◽  
Young’S Modulus ◽  
Experimental Results ◽  
Molecular Weights ◽  
Rubber Specimen ◽  
A Value ◽  
Fractional Increase ◽  
Internal Agreement ◽  
Good Agreement

Abstract The fractional increase in volume accompanying the isothermal extension of soft gum rubber was measured for four rubber samples at mean extensions of 14, 33, and 51%. The chain molecular weights Mc of the four samples were 5500, 5100, 4400, and 3000, with an estimated uncertainty of about 10% in each value of Mc. The observed fractional increase in volume ranged from 3.2×10−5 to 142×10−5, the latter value being observed for the sample of lowest chain molecular weight and at the extension of 51%. The experimental results for each sample have been represented by theoretical curves based on Gee's expression for the fractional increase in volume as a function of the sample extension. The theoretical curves exhibit good agreement with those of Gee, Stern, and Treloar. The process of fitting the theoretical curves to the experimental points constituted a determination of Young's modulus E for each rubber specimen. As a check on the experimental results, and also on the theory employed, determinations of E were also made by two additional methods, namely, from rough stess-strain curves, and from the relation E=3γρRT/Mc. With one exception, the internal agreement between the three determinations of E for the four different samples was satisfactory. The exception noted can probably be ascribed to the use of too small a value of Mc for the sample of lowest chain molecular weight.

Autophagy of mitochondria in rat liver assessed by immunogold procedures.

1988 ◽  
Vol 36 (11) ◽  
pp. 1433-1440 ◽  
Author(s):  
E Knecht ◽  
A Martinez-Ramón ◽  
S Grisolia
Keyword(s):  
Glutamate Dehydrogenase ◽  
Rat Liver ◽  
Polyclonal Antibodies ◽  
Carbamoyl Phosphate ◽  
Autophagic Vacuoles ◽  
The Difference ◽  
In Vivo Administration ◽  
Phosphate Synthase

Glutamate dehydrogenase and carbamoyl phosphate synthase-I were localized in rat liver by immunogold procedures, using monoclonal and polyclonal antibodies. As expected, there was extensive labeling in mitochondria. Label was also found in lysosomal autophagic vacuoles. When autophagy was stimulated by in vivo administration of the anti-microtubular agent vinblastine we found that: (a) carbamoyl phosphate synthase-I and glutamate dehydrogenase could be found in mitochondria within autophagic vacuoles; (b) the carbamoyl phosphate synthase-I and glutamate dehydrogenase content of the mitochondria sequestered into autophagic vacuoles is the same as that of the nearby "free" mitochondria; and (c) in the whole liver, autophagic vacuoles contain c. 1.5 times more glutamate dehydrogenase than carbamoyl phosphate synthase-I, in contrast to mitochondria which have c. three times more carbamoyl phosphate synthase-I than glutamate dehydrogenase. The latter finding could explain, at least partially, the difference in half-lives of these enzymes.

scholarly journals The regulation of carbamoyl phosphate synthase activity in rat liver mitochondria

1976 ◽  
Vol 154 (2) ◽  
pp. 415-421 ◽  
Author(s):  
J D. McGivan ◽  
N M. Bradford ◽  
J Mendes-Mourão
Keyword(s):  
Protein Content ◽  
Nitrogen Source ◽  
Rat Liver ◽  
Liver Mitochondria ◽  
Rat Liver Mitochondria ◽  
Urea Synthesis ◽  
Carbamoyl Phosphate ◽  
Citrulline Synthesis ◽  
Ornithine Transcarbamoylase ◽  
Phosphate Synthase

The rate at which isolated rat liver mitochondria synthesized citrulline with NH4C1 as nitrogen source was markedly dependent on the protein content of the diet. 2. Citrulline synthesis was not rate-limited by substrate concentration, substrate transport or ornithine transcarbamoylase activity under the conditions used. 3. The intramitochondrial content of an activator of carbamoyl phosphate synthase, assumed to be N-acetyl-glutamate, varied markedly with dietary protein content. The variation in the concentration of this activator was sufficient to account for the observed variation in the rates of citrulline synthesis if this synthesis were rate-limited by the activity of carbamoyl phosphate synthase. 4. The rates of urea formation from NH4Cl as nitrogen source in isolated liver cells showed variations in response to diet that closely paralleled the variations in the rates of citrulline synthesis observed in isolated mitochondria. 5. These results are consistent with the postulate that when NH4Cl plus ornithine are present in an excess, the rate of urea synthesis is regulated at the level of carbamoyl phosphate synthase activity.

scholarly journals Purification of membrane-bound galactosyltransferase from rat liver microsomal fractions

1977 ◽  
Vol 164 (3) ◽  
pp. 541-547 ◽  
Author(s):  
Ian H. Fraser ◽  
Sailen Mookerjea
Keyword(s):  
Molecular Weight ◽  
Rat Liver ◽  
Column Chromatography ◽  
High Speed ◽  
Affinity Column ◽  
High Molecular Weight Fraction ◽  
Serum Enzyme ◽  
Membrane Enzyme ◽  
Membrane Bound ◽  
High Speed Supernatant

1. Rat liver microsomal preparations incubated in 1% Triton X-100 at 37°C for 1h released about 60% of the membrane-bound UDP-galactose–glycoprotein galactosyltransferase (EC 2.4.1.22) into a high-speed supernatant. The supernatant galactosyltransferase which was solubilized but not purified by this treatment had a higher molecular weight than the serum enzyme as shown by Sephadex G-100 column chromatography. 2. The galactosyltransferase present in the high-speed supernatant was purified 680-fold by an affinity-column-chromatographic technique by using a column of activated Sepharose 4B coupled with α-lactalbumin. The galactosyltransferase ran as a single band on polyacrylamide gels and contained no sialyltransferase, N-acetylglucosaminyltransferase or UDP-galactose pyrophosphatase activities. 3. The purified membrane enzyme had properties similar to serum galactosyltransferase. It had an absolute requirement for Mn2+ that could not be replaced by Ca2+, Mg2+, Zn2+ or Co2+, and was active over a wide pH range (6–8) with a pH optimum of 6.5. The apparent Km for UDP-galactose was 10.8μm. The protein α-lactalbumin modified the enzyme to a lactose synthetase by increasing substrate specificity for glucose in preference to N-acetylglucosamine and fetuin depleted of sialic acid and galactose. 4. The molecular weight of the membrane enzyme was 65000–70000, similar to that of the purified serum enzyme. Amino acid analyses of the two proteins were similar but not identical. 5. Sephadex G-100 column chromatography of the purified membrane enzyme showed a small peak (2–5%) of higher molecular weight than the purified serum enzyme. Inclusion of 1mm-ε-aminohexanoic acid in the isolation procedures increased this peak to as much as 30% of the total enzyme recovered. Increasing the ε-aminohexanoic acid concentration to 100mm resulted in no further increase in this high-molecular-weight fraction.

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