scholarly journals Molecular weights and metabolism of rat brain proteins

1970 ◽  
Vol 116 (4) ◽  
pp. 745-753 ◽  
Author(s):  
R. Vrba ◽  
Wendy Cannon
Keyword(s):  
Sodium Chloride ◽  
High Speed ◽  
Gel Filtration ◽  
Specific Radioactivity ◽  
Sodium Deoxycholate ◽  
Dynamic State ◽  
Brain Proteins ◽  
Molecular Weights ◽  
Protein Molecules ◽  
The Brain

1. Rats were injected with [U-14C]glucose and after various intervals extracts of whole brain proteins (and in some cases proteins from liver, blood and heart) were prepared by high-speed centrifugation of homogenates in 0.9% sodium chloride or 0.5% sodium deoxycholate. 2. The extracts were subjected to gel filtration on columns of Sephadex G-200 equilibrated with 0.9% sodium chloride or 0.5% sodium deoxycholate. 3. Extracts prepared with both solvents displayed on gel filtration a continuous range of proteins of approximate molecular weights ranging from less than 2×104 to more than 8×105. 4. The relative amount of the large proteins (mol.wt.>8×105) was conspicuously higher in brain and liver than in blood. 5. At 15min after the injection of [U-14C]glucose the smaller protein molecules (mol.wt.<2×104) were significantly radioactive, whereas no 14C could be detected in the larger (mol.wt.>2×104) protein molecules. The labelling of all protein samples was similar within 4h after injection of [U-14C]glucose. Fractionation of brain proteins into distinctly different groups by the methods used in the present work yielded protein samples with a specific radioactivity comparable with that of total brain protein. 6. No evidence could be obtained by the methods used in the present and previous work to indicate the presence of a significant amount of ‘metabolically inert protein’ in the brain. 7. It is concluded that: (a) most or all of the brain proteins are in a dynamic state of equilibrium between continuous catabolism and anabolism; (b) the continuous conversion of glucose into protein is an important part of the maintenance of this equilibrium and of the homoeostasis of brain proteins in vivo.

scholarly journals Active-site determinations on forms of mammalian brain and eel acetylcholinesterase

1976 ◽  
Vol 157 (1) ◽  
pp. 69-76 ◽  
Author(s):  
M A Gordon ◽  
S L Chan ◽  
A J Trevor
Keyword(s):  
Active Site ◽  
Active Sites ◽  
Gel Filtration ◽  
Mammalian Brain ◽  
Vacuum Filtration ◽  
Molecular Weights ◽  
Electric Eel ◽  
Gradient Gel Electrophoresis ◽  
The Brain ◽  
Brain Acetylcholinesterase

Three forms of brain acetylcholinesterase were purified from bovine caudate-nucleus tissue and determined by calibrated gel filtration to have mol.wts. of approx. 120 000 (C), 230 000 (B) and 330 000 (A). [3H]Di-isopropyl phosphorofluoridate (isopropyl moiety labelled) was purified from commercial preparations and its concentration estimated by an enzyme-titration procedure. Brain acetylcholinesterase preparations and enzyme from eel electric tissue were allowed to react with [3H]di-isopropyl phosphorofluridate in phosphate buffer until enzyme activity was inhibited by 98%. Excess of [3H]di-isopropyl phosphorofluoridate that had not reacted was separated from the labelled enzyme protein by gel filtration, or by vacuum filtration or by extensive dialysis. The specificity of active-site labelling was confirmed by use of the enzyme reactivator, pyridine 2-aldoxime. The forms of brain acetylcholinesterase were calculted to contain approximately two (C) four (B) and six (A) active sites per molecule respectively. Acetylcholinesterase (mol.wt. 250 000) from electric-eel tissue was estimated to contain two active sites per molecule. Gradient-gel electrophoresis was used to confirm the estimation of molecular weights of brain acetylcholinesterase forms made by gel filtration. Under the conditions of electrophoresis acetylcholinesterase form A was stable, but form B was converted into a species of approx. 120 000 mol. wt. Similarly, form C of the brain enzyme was converted into a 60 000-mol.wt. form during electrophoresis. These results are in general accord with the suggestion that the multiple forms of brain acetylcholinesterase may be related to the aggregation of a single low-molecular-weight species.

Two types of xylanases of alkalophilic Bacillus sp. No. C-125

1985 ◽  
Vol 31 (6) ◽  
pp. 538-542 ◽  
Author(s):  
H. Honda ◽  
T. Kudo ◽  
Y. Ikura ◽  
K. Horikoshi
Keyword(s):  
Ammonium Sulfate ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Bacillus Sp ◽  
Ammonium Sulfate Precipitation ◽  
Molecular Weights ◽  
Protein Molecules ◽  
Activity Curve ◽  
Alkalophilic Bacillus ◽  
Hydrolysis Of

One alkalophilic Bacillus sp. strain C-125 (FERM No. 7344) was isolated from soil. From this organism, two types of xylanases, designated xylanase A and xylanase N, were purified by an ammonium sulfate precipitation followed by Biogel P-30 gel filtration, DEAE-cellulose chromatography, and Sephadex G-75 gel filtration. The molecular weights of xylanase A and N were estimated as 43 000 and 16 000, respectively. Immunological experiments indicated that xylanase A and xylanase N were entirely different protein molecules. Xylanase N was most active at pH 6.0–7.0, but xylanase A had a very broad pH activity curve (pH 6–10) and was still active even at pH 12.0. The maximum hydrolysis of xylan by the enzymes was about 25%. Both enzymes split xylan and yielded xylobiose and higher oligosaccharides but could hydrolyze neither xylobiose nor xylotriose. Trans xylosidation activities were detected in both enzymes.

Salt tolerance in the halophyte Suaeda maritima : some properties of malate dehydrogenase

1976 ◽  
Vol 273 (927) ◽  
pp. 523-540 ◽  
Keyword(s):  
Sodium Chloride ◽  
Salt Tolerance ◽  
Molecular Mass ◽  
Malate Dehydrogenase ◽  
High Speed ◽  
Gel Filtration ◽  
Supernatant Fraction ◽  
Suaeda Maritima ◽  
Substrate Concentrations ◽  
Mass Form

Malate dehydrogenase activity in a high speed supernatant fraction prepared from homogenates of the shoots of the halophyte Suaeda maritima was separated into two fractions by gel filtration. Sodium chloride stimulated the activity of the larger molecular mass form of the enzyme to a greater extent than that of the smaller molecular mass form : the latter constituted the bulk of the activity. The degree of stimulation by sodium chloride increased with increasing substrate concentrations. Differences between the properties of the enzymes isolated from plants grown with and without sodium chloride are described. The results obtained with the halophyte enzyme were very similar to those obtained with the glycophyte Pisum sativum . Maximum activation of enzyme activity was obtained with a salt concentration of 50 mol m -3 in both species and at higher concentrations activity was inhibited. The results are discussed in relation to the mechanism of salt tolerance in halophytes.

Movement of [U-14C]glucose Carbon into and Subsequent Release from lipids and High-Molecular-Weight Constituents of Rat Brain, Liver, and Heart In Vivo

1972 ◽  
Vol 50 (1) ◽  
pp. 91-105 ◽  
Author(s):  
R. Vrba ◽  
Anna Winter
Keyword(s):  
Amino Acids ◽  
Molecular Weight ◽  
Rat Brain ◽  
High Molecular Weight ◽  
High Speed ◽  
Time Course ◽  
Specific Radioactivity ◽  
Brain Proteins ◽  
Glucose Carbon

After subcutaneous injection of [U-14C]glucose into rats the amount of 14C incorporated in vivo into proteins was always higher than into lipids in brain, liver, and heart. The specific radioactivity of brain proteins was higher than those of liver and heart. Blood-brain comparisons show that protein carbon is derived continuously from glucose in the brain in situ and not as a result of deposition of amino acids or proteins from the circulation. Seventy-two percent of 14C in purified brain protein fractions was found in the amino acids of the hydrolysates of these fractions, mainly in alanine, glutamic, and aspartic acids. Maximum labelling was reached about 4 h after injection of [U-14C]glucose. Elimination of 14C from three classes of brain proteins (high-speed supernatant, particulate deoxycholate extractable, and residual) followed a biphasic time-course. The extent of labelling of, and the rate of elimination of 14C from, the three classes of rat brain proteins were very similar. The fate of 14C in the other investigated tissue fractions of brain, liver, and heart was compared with the fate of 14C in brain proteins.The results lend further support to the previously published suggestion that: (a) brain does not contain appreciable amounts of metabolically inert proteins or of proteins with turnover rates significantly higher than the mean for the bulk of brain proteins; (b) glucose carbon participates at a different rate and to a different extent in the metabolism of high-molecular-weight constituents of brain as compared to liver, heart, and plasma proteins; (c) the continuous conversion of glucose carbon into protein is an important part of the maintenance of the homeostasis of tissue proteins in vivo.

High-Speed Lens-Free Holographic Sensing of Protein Molecules Using Quantitative Agglutination Assays

ACS Sensors ◽  
2021 ◽  
Vol 6 (3) ◽  
pp. 1208-1217
Author(s):  
Zhen Xiong ◽  
Colin J. Potter ◽  
Euan McLeod
Keyword(s):  
High Speed ◽  
Protein Molecules

scholarly journals Purification and characterization of fat body lipase from the greater wax moth, Galleria mellonella (Lepidoptera: Pyralidae)

2019 ◽  
Vol 81 (1) ◽  
Author(s):  
Rahma R. Z. Mahdy ◽  
Shaimaa A. Mo’men ◽  
Marah M. Abd El-Bar ◽  
Emad M. S. Barakat
Keyword(s):  
Metal Ions ◽  
Kinetic Parameters ◽  
Biochemical Characterization ◽  
Galleria Mellonella ◽  
Fat Body ◽  
Specific Activity ◽  
Larval Instar ◽  
Gel Filtration ◽  
Molecular Weights

Abstract Background Insect lipid mobilization and transport are currently under research, especially lipases and lipophorin because of their roles in the production of energy and lipid transport at a flying activity. The present study has been conducted to purify intracellular fat body lipase for the first time, from the last larval instar of Galleria mellonella. Results Purification methods by combination of ammonium sulfate [(NH4)2SO4] precipitation and gel filtration using Sephadex G-100 demonstrated that the amount of protein and the specific activity of fat body lipase were 0.008633 ± 0.000551 mg/ml and 1.5754 ± 0.1042 μmol/min/mg protein, respectively, with a 98.9 fold purity and recovery of 50.81%. Hence, the sephadex G-100 step was more effective in the purification process. SDS-PAGE and zymogram revealed that fat body lipase showed two monomers with molecular weights of 178.8 and 62.6 kDa. Furthermore, biochemical characterization of fat body lipase was carried out through testing its activities against several factors, such as different temperatures, pH ranges, metal ions, and inhibitors ending by determination of their kinetic parameters with the use of p-nitrophenyl butyrate (PNPB) as a substrate. The highest activities of enzyme were determined at the temperature ranges of 35–37 °C and 37–40 °C and pH ranges of 7–9 and 7–10. The partially purified enzyme showed significant stimulation by Ca2+, K+, and Na+ metal ions indicating that fat body lipase is metalloproteinase. Lipase activity was strongly inhibited by some inhibitors; phenylmethylsulfonyl fluoride (PMSF), ethylene-diaminetetractic acid (EDTA), and ethylene glycoltetraacetic acid (EGTA) providing evidence of the presence of serine residue and activation of enzymes by metal ions. Kinetic parameters were 0.316 Umg− 1 Vmax and 301.95 mM Km. Conclusion Considering the purification of fat body lipase from larvae and the usage of some inhibitors especially ion chelating agents, it is suggested to develop a successful control of Galleria mellonella in near future by using lipase inhibitors.

High-Speed Atomic Force Microscopy for Studying the Dynamic Behavior of Protein Molecules at Work

2006 ◽  
Vol 45 (3B) ◽  
pp. 1897-1903 ◽  
Author(s):  
Toshio Ando ◽  
Takayuki Uchihashi ◽  
Noriyuki Kodera ◽  
Atsushi Miyagi ◽  
Ryo Nakakita ◽  
...  
Keyword(s):  
Atomic Force Microscopy ◽  
Dynamic Behavior ◽  
High Speed ◽  
Force Microscopy ◽  
Atomic Force ◽  
Protein Molecules

An Experimental Investigation of the Auto-Ignition Properties of Two C5 Esters: Methyl Butanoate and Butyl Methanoate

2007 ◽  
Author(s):  
Stephen M. Walton ◽  
Carlos Perez ◽  
Margaret S. Wooldridge
Keyword(s):  
Low Temperature ◽  
High Speed ◽  
Combustion Engine ◽  
Low Temperature Combustion ◽  
Moderate Pressure ◽  
Molecular Weights ◽  
Auto Ignition ◽  
Methyl Butanoate ◽  
Temperature Combustion ◽  
Time Histories

Ignition studies of two small esters were performed using a rapid compression facility (RCF). The esters (methyl butanoate and butyl methanoate) were chosen to have matching molecular weights, and C:H:O ratios, while varying the lengths of the constituent alkyl chains. The effect of functional group size on ignition delay time was investigated using pressure time-histories and high speed digital imaging. The mixtures studied covered a range of conditions relevant to oxygenated fuels and fuel additives, including bio-derived fuels. Low temperature and moderate pressure conditions were selected for study due to their relevance to advanced low temperature combustion strategies, and internal combustion engine conditions. The results are discussed in terms of the reaction pathways affecting the ignition properties.

Effect of extractant and molecular size on the optical and chemical properties of soil humic acids

Soil Research ◽  
1969 ◽  
Vol 7 (3) ◽  
pp. 229 ◽  
Author(s):  
JHA Butler ◽  
JN Ladd
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Humic Acids ◽  
Gel Filtration ◽  
Chemical Properties ◽  
Molecular Size ◽  
Carboxyl Content ◽  
Peptide Bonds ◽  
Molecular Weights ◽  
Amino Acid Contents

Humic acids extracted from soil with sodium pyrophosphate have greater proportions of lower molecular weight material, less acid-hydrolysable amino acid nitrogen contents, but greater carboxyl contents and extinction values (260 and 450 nm) than humic acids extracted subsequently from the same sample with alkali. Humic acids extracted with alkali from fresh soil samples have intermediate values. Extinction values at 260 nm are directly correlated with carboxyl contents for a given soil. Different crop histories have no significant effect on the measured properties of the extracted humic acids. An alkali-extracted humic acid has been fractionated by gel filtration into seven fractions of different nominal molecular weight ranges. As the molecular weights of the fractions increase, both aliphatic C-H (based on infrared absorption at 2900 cm-1) and acid-hydrolysable amino acid contents increase, whereas extinction values at 260 nm and carboxyl contents decrease. The infrared spectra of the high molecular weight fractions have peaks at 1650 and 1510 cm-1 which correlate with acid-hydrolysable amino acid contents and which correspond to amide I and II bands of peptide bonds. Alkaline hydrolysis to split peptide bonds eliminates both these peaks. The spectra also have peaks at 1720 and 1210 cm-1 which correlate with the carboxyl content.

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