Detection of basic proteins and low molecular weight peptides in polyacrylamide gels by formaldehyde fixation

Steck, Germaine; Leuthard, Peter; Bürk, R. R.

doi:10.1016/0003-2697(80)90486-8

Cited by 174 publications

(59 citation statements)

References 5 publications

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“…The slab gel was then fixed for 1 h with an aqueous solution composed of 25% ethanol and 14% formaldehyde [17]. Staining and destaining were performed conventionally using Coomassie brilliant blue.…”

Section: Methodsmentioning

confidence: 99%

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Selective labeling of beef heart cytochrome oxidase subunit III with eosin‐5‐maleimide

Müller

Azzi

1985

FEBS Letters

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Cytochrome c oxidase has been isolated from beef heart mitochondria and labeled with the fluorochrome eosin-5-maleimide (EMA) after pretreatment with mersalyl. On SDS-polyacrylamide gels, EMA fluorescence and absorption occurred at a single band corresponding to subunit III. Since only Cys 115 of the two cysteinyl residues of subunit III had been shown to be reactive towards water-soluble SH-reagents, it was concluded that this residue was the one labeled by EMA. The EMA/enzyme ratio was about 1. Gel filtration experiments have shown that upon treatment with dicyclohexylcarbodiimide, subunit III was loosened from the complex; this result suggests that the inhibitory effect of dicyclohexylcarbodiimide on the H+-translocation activity may be related to such a phenomenon. Cytochrome oxidase Subunit III Eosin-5-maleimide

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Section: Methodsmentioning

confidence: 99%

“…Gel permeation was performed on Fractogel TSK, HW-55(S) at 5.4 ml/h in a 1 x 30 cm column and fractions were collected each 15 min after the application of the sample (4-5 nmol enzyme). Protein content was determined using the method of Lowry et al [17].…”

Section: Methodsmentioning

confidence: 99%

Selective labeling of beef heart cytochrome oxidase subunit III with eosin‐5‐maleimide

Müller

Azzi

1985

FEBS Letters

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“…Digestion products were separated by SDS-PAGE using Tris-borate as running buffer (5). After electrophoresis, the gel was formaldehyde-fixed and stained with Coomassie brilliant blue R-250 (25).…”

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confidence: 99%

Purification and Characterization of Arcelin Seed Protein from Common Bean

Osborn¹,

Burow²,

Bliss³

1988

Plant Physiol.

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Arcelin, a seed protein originally discovered in wild bean accessions, was purified, characterized, and compared to phaseolin, the major seed protein of common bean, and to phytohemagglutinin (PHA), the major bean seed lectin. Arcelin and PHA has several characteristics in common.Both were glycoproteins having similar subunit M, deglycosylated Mn and amino acid compositions. The two proteins were related antigenically and they had the same developmental timing of accumulation. Arcelin also had some hemagglutinating activity, a characteristic associated with lectins. However, several features distinguished arcelin from PHA. Arcelin had a more basic isoelectric point than PHA, greater numbers of basic amino acid residues, additional cysteine residues, and one methionine residue, which PHA lacks. Native PHA protein is a tetramer of subunits, and although a small component of native arcelin protein was also tetrameric, most of the arcelin preparation was dimeric. The hemagglutinating activity ofarcelin was specific only for some pronase-treated erythrocytes. It did not agglutinate native erythrocytes, nor did it bind to thyroglobulin or fetuin aff'mity resins as did PHA. Although arcelin has lectin-like properties, we believe the distinctions between arcelin and PHA warrant the designation of arcelin as a unique bean seed protein.was co-dominant with respect to each other and dominant with respect to alleles for the absence of arcelin. Genes controlling arcelin expression were tightly linked (<0.3% recombination) to those controlling PHA expression. The presence of arcelin in wild bean accessions was correlated with high levels of resistance to two bruchid beetle species (19,24).Although arcelin protein has a different electrophoretic mobility than other major seed proteins, it is not known whether arcelins represent a truly distinct class of seed proteins. The relationship of arcelin to other seed proteins is particularly interesting in light of the tight linkage between arcelin and PHA genes. In this study, we report on the purification and characteriz,tion of the arcelin-1 variant and compare its properties to those of PHA and phaseolin seed proteins. The major seed storage proteins of cultivated common bean (Phaseolus vulgaris) are phaseolin and PHA,2 also referred to as bean lectin. In several wild bean accessions, a third important protein fraction has been observed which had not been described previously in bean cultivars (19,23). This fraction was discovered as a major seed protein having a different electrophoretic mobility than either phaseolin or PHA with which it occurred in seeds of some wild beans. Four electrophoretic variants of this protein were observed which consisted of distinct groups of polypeptides having similar mobilities on two-dimensional gels. This protein was named arcelin and the variants were designated arcelin-1, 2, 3, and 4.There has been only limited characterization of arcelin, but the protein and its genetic control have several interesting features. Arcelin was shown to o...

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“…Samples dissolved in 8 M urea, 0.9 M acetic acid, 0.5 M 2-mercaptoethanol were run in slab gels (140 x 200 x 0.75 mm) containing 17% acrylamide and 6.25 M urea. Gels were stained with Coomassie brilliant blue R 250 as in [16].…”

Section: Analytical Gel Electrophoresismentioning

confidence: 99%

Nuclear basic protein transition during sperm differentiation. Amino acid sequence of a spermatid-specific protein from the dog-fish Scylliorhinus caniculus

Chauvière¹,

Martinage²,

Briand³

et al. 1987

Eur J Biochem

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During dog-fish spermatogenesis, chromatin undergoes a continuous processing which involves two basic protein transitions : the first from somatic-type histones to spermatid-specific proteins and the second leading to protamines. Two spermatid-specific proteins S1 and S2 were isolated from nuclei of spermatid-enriched testis zone and the amino acid sequence of S1 has been determined. S1 contains 87 amino acids and has a molecular mass of 11 179 Da. It is mainly characterized by a high content of basic residues (45%) and the presence of one residue of cysteine. Its primary structure shows that the N-terminal half is highly basic while the hydrophobic residues are preferentially localized in the C-terminal region. Three forms of S1 are present in testis which correspond to di-, mono-and nonphosphorylated molecules. This spermatid-specific protein shares no common structural feature with either histones and dogfish protamines or rat spermatid-specific protein which has been previously described.In most animal species, spermatogenesis is characterized by dramatic morphological and biological changes in nuclear organization. In sperm nuclei, four types of DNA-associated proteins have been described [l]: (a) somatic-type histones or specific histones; (b) proteins with intermediate properties between histones and protamines; (c) low-molecular-mass arginine-rich protamines ; (d) arginine-and cysteine-rich protamines.During sperm differentiation processes, there is generally only one protein transition from histones to sperm-specific proteins; however, in some species such as Eutherian mammals [2, 31 and dog-fish [4, 51, specific and often heterogeneous basic proteins occur transiently during spermiogenesis before the setting-down of the protamines on the DNA (for review, see [6]). In somatic cells, the conservajion of histone primary structures is accompanied by a very conserved nucleosomal organization of chromatin throughout evolution. In contrast, in germ cells, the wide diversity of the nuclear basic protein equipment may lead to many possibilities of DNA packaging.Therefore, it is of great interest to compare, both within one species and between different species, the structures of the various proteins which package DNA during spermatogenesis. Dog-fish is an attractive model both for practical and biological reasons. First, during dog-fish spermatogenesis, the development of germ cells is synchronous within follicles and transverse sections of testis show a zonation in follicle maturaCorrespondence to M.

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Detection of basic proteins and low molecular weight peptides in polyacrylamide gels by formaldehyde fixation

Cited by 174 publications

References 5 publications

Selective labeling of beef heart cytochrome oxidase subunit III with eosin‐5‐maleimide

Selective labeling of beef heart cytochrome oxidase subunit III with eosin‐5‐maleimide

Purification and Characterization of Arcelin Seed Protein from Common Bean

Nuclear basic protein transition during sperm differentiation. Amino acid sequence of a spermatid-specific protein from the dog-fish Scylliorhinus caniculus

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