Isolation of Labeled Lipoprotein from Escherichia coli and Proteus mirabilis after Incubation with [14C]Penicillin

1985 ◽  
Vol 40 (5-6) ◽  
pp. 415-420 ◽  
Author(s):  
Gerhard Gruner ◽  
Monier H. Tadros ◽  
Roland Plapp
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Proteus Mirabilis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Apparent Molecular Weight ◽  
Penicillin G ◽  
Free Form ◽  
E Coli

Abstract [14C]penicillin binding experiments and membrane analysis were carried out with cell envelope preparations from Escherichia coli and Proteus mirabilis. After incubation with [14C] penicillin G labeled free lipoprotein could be identified. The analysis of the isolated lipoprotein by SDS polyacrylamide gel electrophoresis indicates that there is only one protein with an apparent molecular weight of 7000. The amino acid composition of isolated labeled free lipoprotein from E. coli was identical to the lipoprotein already found in E. coli. It is a point of interest that the amino acid composition of the isolated labeled free lipoprotein from P. mirabilis D52 differs from that found in other mutants of this strain. The free form of lipoprotein from P. mirabilis D52 is composed of 61 amino acids and has glycine, phenylalanine and proline as specific components.

scholarly journals Separation and partial characterization of guinea-pig caseins

1978 ◽  
Vol 173 (2) ◽  
pp. 633-641 ◽  
Author(s):  
R K Craig ◽  
D McIlreavy ◽  
R L Hall
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Guinea Pig ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Exchange Chromatography

1. Guinea-pig caseins A, B and C were purified free of each other by a combination of ion-exchange chromatography and gel filtration. 2. Determination of the amino acid composition showed all three caseins to contain a high proportion of proline and glutamic acid, but no cysteine. This apart, the amino acid composition of the three caseins was markedly different, though calculated divergence values suggest that some homology may exist between caseins A and B. Molecular-weight estimates based on amino acid composition were in good agreement with those based on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. 3. N-Terminal analysis showed lysine, methionine and lysine to be the N-terminal residues of caseins A, B and C respectively. 4. Two-dimensional separation of tryptic digests revealed a distinctive pattern for each casein. 5. All caseins were shown to be phosphoproteins. The casein C preparation also contained significant amounts of sialic acid, neutral and amino sugars. 6. The results suggest that each casein represents a separate gene product, and that the low-molecular-weight proteins are not the result of a post-translational cleavage of the largest. All were distinctly different from the whey protein alpha-lactalbumin.

scholarly journals Studies on the subunits of human myeloperoxidase

1984 ◽  
Vol 222 (3) ◽  
pp. 701-709 ◽  
Author(s):  
R L Olsen ◽  
C Little
Keyword(s):  
Heat Treatment ◽  
Amino Acid ◽  
Molecular Mass ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Additional Species ◽  
Main Protein

The subunit composition of human myeloperoxidase was studied with the use of sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and gel filtration. The subunit pattern observed depended on the manner in which the enzyme was treated before analysis. Reduction before heat treatment in detergent led to two main protein species (Mr 57 000 and 10 500), whereas reduction during or after heat treatment yielded an additional species of Mr 39 000. Heating without any reductive pretreatment yielded the 39 000-Mr form as the major electrophoretic species. Carbohydrate staining showed large amounts of sugar on the 57 000-Mr species and little on the 10 500-Mr form. Significant amounts of haem were associated with this latter subunit. Haem also seemed to be associated with the 57 000-Mr form but not with the 39 000-Mr one. These three subunit forms were isolated and their amino acid composition analysed. The 57 000-Mr and 39 000-Mr forms had very similar amino acid composition and yielded an apparently identical collection of fragments on incubation with CNBr. Once separated, the subunits could not be interconverted. Generally, minor amounts of other molecular-mass forms were observed. The nature of the various molecular-mass forms originating from myeloperoxidase is discussed.

scholarly journals Purification and biochemical characterization of three major acidic proteins (BSP-A1, BSP-A2 and BSP-A3) from bovine seminal plasma

1987 ◽  
Vol 241 (3) ◽  
pp. 685-692 ◽  
Author(s):  
P Manjunath ◽  
M R Sairam
Keyword(s):  
Amino Acid ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Seminal Plasma ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Polyacrylamide Gel ◽  
Terminal Residue ◽  
Acidic Proteins

Three major acidic proteins of bovine seminal plasma, BSP-A1, BSP-A2 and BSP-A3, were purified to homogeneity, by employing fast protein liquid chromatography, gel filtration and h.p.l.c. The proteins were purified on the basis of their stimulatory effect on the basal release of gonadotropins by rat anterior-pituitary cells in culture. All three proteins migrated as distinct single bands in the presence or absence of 2-mercaptoethanol in SDS/polyacrylamide-gel electrophoresis. Their Mr values were estimated to be between 15,000 and 16,500 by SDS/polyacrylamide-gel electrophoresis. Similar Mr estimates were obtained when they were subjected to gel filtration on a calibrated column of Sephadex G-75 equilibrated in 0.05 M-acetic acid, pH 3.0. However, BSP-A1 and BSP-A2 were eluted as aggregated molecules (Mr 60,000-120,000) during gel filtration on Sephadex G-200 equilibrated in 0.05 M-NH4HCO3, pH 8.5, or phosphate buffer, pH 7.0, containing 0.15 M-NaCl. In the presence of 8 M-urea both BSP-A1 and BSP-A2 were eluted at positions corresponding to Mr values of 17,000-20,000. BSP-A1 and BSP-A2 had an identical amino acid composition, which differed largely from that of BSP-A3. All three proteins contained aspartic acid as the N-terminal residue, and cysteine was identified as the C-terminal residue. BSP-A1 and BSP-A2 are glycoproteins containing galactosamine, sialic acid and neutral sugars, but BSP-A3 did not contain any covalently attached sugars. Whereas BSP-A2 and BSP-A3 were eluted unadsorbed, BSP-A1 bound to wheat-germ lectin-Sepharose 6MB and could be eluted by the competing sugar N-acetyl-D-glucosamine. Treatment of BSP-A1 and BSP-A2 with trypsin resulted in complete loss of gonadotropin-release activity, but BSP-A3 retained full activity. Antibody raised against BSP-A1 did not cross-react with BSP-A3, or vice versa. All these properties indicated marked structural differences between BSP-A3 and BSP-A1 (or BSP-A2). On the basis of amino acid composition it was concluded that BSP-A1, BSP-A2 and BSP-A3 are the same as the gonadostatins [Esch, Ling, Bohlen, Ying & Guillemin (1983) Biochem. Biophys. Res. Commun. 113, 861-867].

scholarly journals More benign random peptides in E. coli are enriched for similar amino acids as young animal but not plant genes

2020 ◽  
Author(s):  
Luke Kosinski ◽  
Nathan Raul Aviles ◽  
Kevin Gomez ◽  
Joanna Masel
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
De Novo ◽  
Young Animal ◽  
Structural Disorder ◽  
E Coli ◽  
Random Peptide ◽  
Almost All

Proteins are the workhorses of the cell, yet they carry great potential for harm via misfolding and aggregation. Despite the dangers, proteins are sometimes born de novo from non-coding DNA. Proteins are more likely to be born from non-coding regions that produce peptides that do little to no harm when translated than from regions that produce harmful peptides. To investigate which newborn proteins are most likely to "first, do no harm", we estimate fitnesses from an experiment that competed Escherichia coli lineages that each expressed a unique random peptide. A variety of peptide metrics significantly predict lineage fitness, but almost all this predictive power stems from simple amino acid composition. Amino acids that are smaller and that promote intrinsic structural disorder have more benign fitness effects. Our amino acid composition-based predictions also have validity for an independent dataset using small random N-terminal tags. The same amino acids that predict high fitness in E. coli are enriched in young Pfams in animals, but not in plants. To modify Jacques Monod's famous quote, what makes peptides benign in E.coli also makes them benign in elephants, but not in eucalyptus.

scholarly journals The heterogeneity of the non-aggregating proteoglycans of the human intervertebral disc

1987 ◽  
Vol 244 (1) ◽  
pp. 27-33 ◽  
Author(s):  
J L DiFabio ◽  
R H Pearce ◽  
B Caterson ◽  
H Hughes
Keyword(s):  
Amino Acid ◽  
Intervertebral Disc ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Gel Electrophoresis ◽  
Chondroitin Sulphate ◽  
Polyacrylamide Gel Electrophoresis ◽  
Keratan Sulphate ◽  
Volume Analysis ◽  
Molecular Masses

Non-aggregating proteoglycans of differing average hydrodynamic volumes were prepared from nuclei pulposi and anuli fibrosi of three human lumbar spines and characterized by biochemical and immunochemical analyses. The hexose-to-hexuronate and protein-to-hexuronate ratios increased with decreasing hydrodynamic volume. Analysis by composite agarose/polyacrylamide-gel electrophoresis has demonstrated two aggregating subpopulations [McDevitt, Jahnke & Green (1982) Trans. Annu. Meet. Orthop. Res. Soc. 7, 50]. In the present study, electrophoresis of the non-aggregating pools has shown three additional subpopulations, here named bands III, IV and V. The two smallest proteoglycan pools from each tissue contained two and three components respectively. These components were isolated by preparative electrophoresis and analysed. Band III was a proteoglycan richer in keratan sulphate than in chondroitin sulphate; band IV was a proteoglycan richer in chondroitin sulphate than in keratan sulphate; band V contained only chondroitin sulphate. Unsaturated disaccharides prepared from the chondroitin sulphate of all bands were predominantly 6-sulphated, with only 5-15% 4-sulphated. The molecular masses of the chondroitin sulphate and keratan sulphate did not differ between the bands. The amino acid composition of band III differed from that of band IV. Thus three distinct subpopulations of non-aggregating proteoglycan were demonstrated in the human intervertebral disc.

scholarly journals Ethanol Extraction of Basic Proteins From Ejaculated Human Spermatozoa

1979 ◽  
Vol 32 (5) ◽  
pp. 443 ◽  
Author(s):  
Peter French
Keyword(s):  
Amino Acid ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Banding Pattern ◽  
Gel Electrophoresis ◽  
Basic Protein ◽  
Polyacrylamide Gel Electrophoresis ◽  
Ethanol Extraction ◽  
Basic Proteins ◽  
Unconventional Method

A method for the extraction of basic proteins from human ejaculated spermatozoa has been developed It relies on the previously unreported observation that such basic protein is soluble in a solution containing 60% (v/v) ethanol. This unconventional method yields a high percentage of arginine-rich basic protein which is then able to be characterized on the basis of its amino acid composition. This method also allows comparisons to be made between single ejaculates by the banding pattern each displays when subjected to polyacrylamide gel electrophoresis.

scholarly journals The purification and properties of the β-lactamase specified by the resistance factor R-1818 in Escherichia coli and Proteus mirabilis

1971 ◽  
Vol 123 (4) ◽  
pp. 493-500 ◽  
Author(s):  
J. W. Dale ◽  
J. T. Smith
Keyword(s):  
Escherichia Coli ◽  
Proteus Mirabilis ◽  
Gel Electrophoresis ◽  
Specific Activity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Resistance Factor ◽  
Ph Optimum ◽  
E Coli ◽  
R Factor ◽  
Purification And Properties

1. The β-lactamase specified by the R-1818 resistance factor in Escherichia coli was purified 300-fold; the resulting preparation gave a single peak on Sephadex G-100 and a single band on polyacrylamide-gel electrophoresis. 2. The β-lactamase specified by the same R-factor in Proteus mirabilis was purified over 2000-fold, but was still far from pure. The specific activity of this preparation was one-fifth that of the purified enzyme from E. coli. 3. The two enzymes were shown to be identical as regards substrate specificity, pH optimum, Km values and molecular weight. 4. It is suggested that the low β-lactamase activity of extracts of P. mirabilis (R-1818), about 5% of that from E. coli (R-1818) in crude extracts, could be due to inefficient transcription of the R-factor DNA by Proteus RNA polymerase.

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