[49] Reaction of proteins with citraconic anhydride

Atassi, M. Zouhair; Habeeb, A.F.S.A.

doi:10.1016/s0076-6879(72)25053-4

Cited by 189 publications

(86 citation statements)

References 13 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Chain separation of goldfish globin was achieved by chromatography on Sephacryl S200 (30% acetic acid) following denaturation (8 M urea, pH 8.6) and o-chain disulfide polymer formation (oxidized in presence of dehydroascorbic acid) [ 161; for human globin chains by standard procedures [ 171. Aminoethylation [ 181, cyanogen bromide fragmentation [ 193, citraconylation [20] and maleylation [2 l] were performed as indicated. Tryptic digestions (Sigma type XI, TPCK-treated enzyme) were done in 0.2 M N-ethylmorpholine acetate (pH 8.0) using a 2% (w/w) protease ratio at 37°C and terminated by diluting into the starting buffer for the chromatographic separation.…”

Section: Methodsmentioning

confidence: 99%

Microsequence analysis: I. Peptide isolation using high‐performance liquid chromatography

1979

View full text Add to dashboard Cite

Section: Methodsmentioning

confidence: 99%

Microsequence analysis: I. Peptide isolation using high‐performance liquid chromatography

1979

View full text Add to dashboard Cite

“…Citraconylation was executed essentially as described by Atassi and Habeeb [12]. Histone H2Ac1, (30 mg) was dissolved in 2 ml 6 M urea and 4 ml 0.2 M sodium bicarbonate buffer pH 8.5 was added.…”

Section: Fragmentation Of H2a(mentioning

confidence: 99%

The amino acid sequence of wheat histone H2A(1). A core histone with a C-terminal extension

A¹,

Wf²,

C³

1985

Eur J Biochem

View full text Add to dashboard Cite

1. The complete amino acid sequence (145 residues) of histone variant H2A,,, from wheat germ Triticum 2. The sequence of histone variant H2A(,, differs from the homologous calf histone in 61 amino acid positions. aestivum cultivar T 4 has been established from Edman degradation of large overlapping fragments.These differences include an extension of H2A,,, by 19 amino acids at its carboxyl end.Histones in general are considered to play a structural rather than a regulatory role in chromatin [l], however the widespread occurrence of histone variants (isohistones), (for review see [2,31) suggests that variants may modulate the conformation of nucleosomes, and in this way modify their susceptibility to regulatory factors. We have established previously the presence of a number of isohistones of the H2A class in wheat embryos [2, 41. We are reporting now the complete structure of one of these isohistones. This structure constitutes a new histone type in that its C-terminal region is extended beyond the size hitherto considered typical for core histones of that group. MATERIALS AND METHODS Isolation of histone H2A(,,Wheat germ (Triticum aestivum) was obtained from Sasko Mills (Rondebosch, Cape). This material contained at least 80% of the cultivar T 4. Chromatin was isolated essentially according to the method of Johns and Butler [5] with some modifications as described previously [4].Exclusion chromatography of histones was performed at room temperature on a column (4.5 x 80 cm) of Bio-Gel P-60 (100-200 mesh). Approximately 200 mg of total acid-extracted (0.2 M H2S04) histone was dissolved in 10 ml of 6 M urea containing 2% (v/v) 2-mercaptoethanol, loaded onto the column and eluted with 0.02 M HCI (Fig. 1 a).Cation-exchange chromatography was carried out using LKB CM-Trisacryl M equilibrated with 0.05 M sodium acetate adjusted to pH 5.00 with HC1. A linear NaCl gradient was used to elute the H2A(,,-enriched fraction B (Fig. 1 b). Fraction B was then further purified by passage over a BioGel P-60 column equilibrated with 0.02 M HCl, 0.05 M NaCl as described previously [6]. The eluant contained in addition, 10 mM sodium bisulfite, 1 mM PhMeS02F and 1 mM Natosyllysylchloromethane, solubilized first in a small volume of propan-1-01 (Fig. 1 c). The H2A(,,-containing fraction was sometimes recycled over the P-60 column if there was still some contamination evident on gel electrophoresis. Analytical proceduresPolyacrylamide gel electrophoresis (PAGE) employing the detergent Triton X-100 was performed in glass tubes (9 x 0.5 cm) [7, 81.Histone samples were hydrolysed under vacuum for 24 h at 110°C in constant-boiling HCl containing 0.025% (w/v) phenol as antioxidant unless otherwise stated. The analyses were done with either ninhydrin methodology (Beckman 119 automatic analyser) or using hypochlorite and o-phthalaldehyde post-column fluorescence detection (Waters Associates HPLC amino acid analyser [9]).Fragmentation of H2A(,, N-Bromosuccinimide (SucNBr) cleavage [lo] was performed as described previously [ll]. The increase...

show abstract

“…Modijication of lysine residues with citraconic anhydride in order to prevent the cleavage of lysyl bonds was performed essentially according to Atassi and Habeeb [15]. The reaction was performed in distilled water adjusted to pH 8.5 with 5 M NaOH using a Radiometer titrator unit.…”

Section: Enzyme Digestions and Purfication Of Peptidesmentioning

confidence: 99%

Structural Studies on the Four Repetitive Fc-Binding Regions in Protein A from Staphylococcus aureus

Sjödahl¹

1977

Eur J Biochem

121

View full text Add to dashboard Cite

The covalent structures of the four highly homologous Fc-binding regions in protein A, regions D, A, B, and C, have been studied by enzymic fragmentations of previously isolated fragments originating from these regions and subsequent isolation of the generated peptides by ion-exchange chromatography, molecular-sieve chromatography, high-voltage paper electrophoresis and paper chromatography. The complete sequence of region B was elucidated by combining the results of Edman degradations on isolated fragment B peptides with the previously reported N-terminal sequence of the same fragment. Furthermore, Edman degradations of fragment D, A and C peptides differing from the region B sequence provided the structures of subregions not identical to corresponding subregions within region B. Thus, it is possible to propose a highly probable covalent structure for the N-terminal 27 000-molecular-weight portion of protein A responsible for the IgG-Fc-binding activities. However, it was not possible to assign the activities to specific structures within the regions.The sequence data indicate that not only mutual homology between the four regions exists, but also internal homologies within the regions. Furthermore, the data strongly supports the hypothesis of a stepwise gene fusion procedure being involved in the evolution of the protein.

show abstract

[49] Reaction of proteins with citraconic anhydride

Cited by 189 publications

References 13 publications

Microsequence analysis: I. Peptide isolation using high‐performance liquid chromatography

Microsequence analysis: I. Peptide isolation using high‐performance liquid chromatography

The amino acid sequence of wheat histone H2A(1). A core histone with a C-terminal extension

Structural Studies on the Four Repetitive Fc-Binding Regions in Protein A from Staphylococcus aureus

Contact Info

Product

Resources

About